Rice hybrid hr120001

ABSTRACT

A rice hybrid designated HR120001 (ATCC PTA-13290) is disclosed. The invention relates to the seeds of rice hybrid HR120001, to the plants of rice hybrid HR120001 and to methods for producing a rice plant produced by crossing the hybrid HR120001 with itself or another rice plant. The invention further relates to hybrid rice seeds and plants produced by crossing the hybrid HR120001 with another rice plant. This invention further relates to growing and producing blends of rice seeds comprised of seeds of rice hybrid HR120001 with rice seed of one, two, three, four, or more of another rice hybrid, rice variety or rice inbred.

CROSS REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of priority under 35 U.S.C. §119(e)to U.S. Provisional Application No. 61/698,378 filed Sep. 7, 2012. Thedisclosures set forth in the referenced application are incorporatedherein by reference in their entireties, including all information asoriginally submitted to the United States Patent and Trademark Office.

BACKGROUND OF THE INVENTION

The present invention relates to a new and distinctive rice hybriddesignated HR120001. All publications cited in this application areherein incorporated by reference.

Rice is an ancient agricultural crop and is today one of the principalfood crops of the world. There are two cultivated species of rice: Oryzasativa L., the Asian rice, and Oryza glaberrima Steud., the Africanrice. The Asian species constitutes virtually all of the world'scultivated rice and is the species grown in the United States. Threemajor rice producing regions exist in the United States: the MississippiDelta (Arkansas, Mississippi, northeast Louisiana, southeast Missouri),the Gulf Coast (southwest Louisiana, southeast Texas), and the CentralValleys of California.

Rice production in the United States can be broadly categorized aseither dry-seeded or water-seeded. In the dry-seeded system, rice issown into a well-prepared seed bed with a grain drill or by broadcastingthe seed and incorporating it with a disk or harrow. Moisture for seedgermination is from irrigation or rainfall. Another method of plantingby the dry-seeded system is to broadcast the seed by airplane into aflooded field, then promptly drain the water from the field. For thedry-seeded system, when the plants have reached sufficient size (four-to five-leaf stage), a shallow permanent flood of water 5 to 16 cm deepis applied to the field until the rice approaches maturity. Rice isgrown on flooded soils to optimize grain yields. Heavy clay soils orsilt loam soils with hard pan layers about 30 cm below the surface aretypical rice-producing soils because they minimize water losses due topercolation.

In the water-seeded system, rice seed is soaked for 12 to 36 hours toinitiate germination, and the seed is broadcast by airplane into ashallow-flooded field. Water may be drained from the field for a shortperiod of time to enhance seedling establishment or the seedlings may beallowed to emerge through the shallow flood. In either case, a shallowflood is maintained until the rice approaches maturity. For both thedry-seeded and water-seeded production systems, the rice is harvestedwith large combines 2 to 3 weeks after draining.

Rice in the United States is classified into three primary market typesby grain size and shape as: long-grain, medium grain and short-grain.Typical U.S. long-grain rice cooks dry and fluffy when steamed orboiled, whereas medium- and short-grain rice cooks moist and sticky.Long-grain cultivars have been traditionally grown in the southernstates and generally receive higher market prices.

Although specific breeding objectives vary somewhat in the differentregions, increasing yield is a primary objective in all programs. Grainyield of rice is determined by the number of panicles per unit area, thenumber of fertile florets per panicle, and grain weight per fertilefloret. Increases in any or all of these yield components provide amechanism to obtain higher yields. Heritable variation exists for all ofthese components, and breeders may directly or indirectly select forincreases in any of them.

There are numerous steps in the development of any novel, desirablecultivar. Plant breeding begins with the analysis and definition ofproblems and weaknesses of the current cultivars, followed by theestablishment of program goals, and the definition of specific breedingobjectives. The next step is selection of parental lines that possessthe traits required to meet the program goals. The goal is to combine ina single cultivar an improved combination of desirable traits from theparental sources. These important traits may include higher yield,resistance to diseases and insects, better stems and roots, tolerance tolow temperatures, better agronomic characteristics, and grain quality.

The goal of rice plant breeding is to develop new, unique, and superiorrice cultivars and hybrids. The breeder initially selects and crossestwo or more parental lines, followed by selection among the many newgenetic combinations. The breeder can theoretically generate billions ofnew and different genetic combinations via crossing. The breeder has nodirect control at the cellular level; therefore, two breeders will neverdevelop the same line, or even very similar lines, having the same ricetraits.

Choice of breeding methods to select for the improved combination oftraits depends on the mode of plant reproduction, the heritability ofthe trait being improved, and the type of cultivar used commercially(e.g., F₁ hybrid cultivar, pure line cultivar, etc.). For highlyheritable traits, a choice of superior individual plants evaluated at asingle location will be effective, whereas for traits with lowheritability, selection should be based on mean values obtained fromreplicated evaluations of families of related plants. Popular selectionmethods include pedigree selection, backcross selection, and single seedselection, or a combination of these methods.

Pedigree breeding is used commonly for the improvement ofself-pollinating crops such as rice. Two parents which possessfavorable, complementary traits are crossed to produce an F₁. One orboth parents may themselves represent an F₁ from a previous cross.Subsequently a segregating population is produced, growing the seedsresulting from selfing one or several F₁s if the two parents are purelines or by directly growing the seed resulting from the initial crossif at least one of the parents is an F₁. Selection of the bestindividuals may begin in the first segregating population or F₂; then,beginning in the F₃, the best individuals in the best families areselected. Replicated testing of families can begin in the F₄ generationto improve the effectiveness of selection for traits with lowheritability. At an advanced stage of inbreeding (i.e., F₆ and F₇), thebest lines or mixtures of phenotypically similar lines are tested forpotential release as new parental lines.

Mass and recurrent selections can be used to improve populations ofeither self- or cross-pollinating crops. A genetically variablepopulation of heterozygous individuals is either identified or createdby intercrossing several different parents. The best plants are selectedbased on individual superiority, outstanding progeny, or excellentcombining ability. The selected plants are intercrossed to produce a newpopulation in which further cycles of selection are continued.

Backcross breeding has been used to transfer genes for a highlyheritable trait into a desirable homozygous cultivar or inbred linewhich is the recurrent parent. The source of the trait to be transferredis called the donor parent. The resulting plant is expected to have theattributes of the recurrent parent (e.g., cultivar) and the desirabletrait transferred from the donor parent. After the initial cross,individuals possessing the phenotype of the donor parent are selectedand repeatedly crossed (backcrossed) to the recurrent parent.

The single-seed descent procedure in the strict sense refers to plantinga segregating population, harvesting a sample of one seed per plant, andusing the one-seed sample to plant the next generation. When thepopulation has been advanced from the F₂ to the desired level ofinbreeding, the plants from which lines are derived will each trace todifferent F₂ individuals. The number of plants in a population declineseach generation due to failure of some seeds to germinate or some plantsto produce at least one seed. As a result, not all of the F₂ plantsoriginally sampled in the population will be represented by a progenywhen generation advance is completed.

In a multiple-seed procedure, rice breeders commonly harvest one or moreseeds from each plant in a population and thresh them together to form abulk. Part of the bulk is used to plant the next generation and part isput in reserve. The procedure has been referred to as modifiedsingle-seed descent or the pod-bulk technique.

The multiple-seed procedure has been used to save labor at harvest. Itis considerably faster to thresh panicles with a machine than to removeone seed from each by hand for the single-seed procedure. Themultiple-seed procedure also makes it possible to plant the same numberof seeds of a population each generation of inbreeding. Enough seeds areharvested to make up for those plants that did not germinate or produceseed.

Descriptions of other breeding methods that are commonly used fordifferent traits and crops can be found in one of several referencebooks (e.g., Allard, R. W. et al. Principles of Plant Breeding (1999);Agrawal, R. L. Fundamentals of Plant Breeding and Hybrid Seed Production(1998); Schlegel, R. H. J. Encyclopedic Dictionary of plant Breeding andRelated Subjects (2003); Fehr, W. R. et al. Principles of CultivarDevelopment—Theory and Technique (1987)).

Each breeding program should include a periodic, objective evaluation ofthe efficiency of the breeding procedure. Evaluation criteria varydepending on the goal and objectives, but should include gain fromselection per year based on comparisons to an appropriate standard,overall value of the advanced breeding lines, and number of successfulcultivars produced per unit of input (e.g., per year, per dollarexpended, etc.).

Promising advanced breeding lines are thoroughly tested and compared toappropriate standards in environments representative of the commercialtarget area(s) for at least three or more years. The best lines arecandidates for new commercial cultivars; those still deficient in a fewtraits may be used as parents to produce new populations for furtherselection.

These processes, which lead to the final step of marketing anddistribution, usually take from 8 to 12 years from the time the firstcross is made and may rely on the development of improved breeding linesas precursors. Therefore, development of new cultivars is atime-consuming process that requires precise forward planning, efficientuse of resources, and a minimum of changes in direction.

A most difficult task is the identification of individuals that aregenetically superior, because for most traits the true genotypic valueis masked by other confounding plant traits or environmental factors.One method of identifying a superior plant is to observe its performancerelative to other experimental plants and to a widely grown standardcultivar. If a single observation is inconclusive, replicatedobservations provide a better estimate of its genetic worth.

Each breeding cycle, the plant breeder selects the germplasm to advanceto the next generation. This germplasm is grown under unique anddifferent geographical, climatic and soil conditions and furtherselections are then made throughout the growing season. The cultivarswhich are developed are unpredictable. This unpredictability is becausethe breeder's selection occurs in unique environments, with no controlat the DNA level (using conventional breeding procedures), and withmillions of different possible genetic combinations being generated. Abreeder of ordinary skill in the art cannot predict the final resultinglines he develops, except possibly in a very gross and general fashion.The same breeder cannot produce the same cultivar twice by using theexact same original parents and the same selection techniques. Thisunpredictability results in the expenditure of large amounts of researchmonies to develop superior new rice cultivars.

Proper testing should detect any major faults and establish the level ofsuperiority or improvement over current cultivars. In addition toshowing superior performance, there must be a demand for a new cultivarthat is compatible with industry standards or which creates a newmarket. The introduction of a new cultivar will incur additional coststo the seed producer, the grower, processor and consumer; for specialadvertising and marketing, altered seed and commercial productionpractices, and new product utilization. The testing preceding release ofa new cultivar should take into consideration research and developmentcosts as well as technical superiority of the final cultivar. Forseed-propagated cultivars, it must be feasible to produce seed easilyand economically.

Rice, Oryza sativa L., is an important and valuable field crop. Thus, acontinuing goal of rice breeders is to develop stable, high yieldingrice cultivars that are agronomically sound. The reasons for this goalare to maximize the amount of grain produced on the land used and tosupply food for both animals and humans. To accomplish this goal, therice breeder must select and develop rice plants that have the traitsthat result in superior cultivars.

The foregoing examples of the related art and limitations relatedtherewith are intended to be illustrative and not exclusive. Otherlimitations of the related art will become apparent to those of skill inthe art upon a reading of the specification.

SUMMARY OF THE INVENTION

The following embodiments and aspects thereof are described inconjunction with systems, tools and methods which are meant to beexemplary and illustrative, not limiting in scope. In variousembodiments, one or more of the above-described problems have beenreduced or eliminated, while other embodiments are directed to otherimprovements.

The present invention relates to a hybrid rice designated HR120001, andseeds and plants derived from the hybrid. The invention also relates tohybrid plants and seeds and any further progeny or descendants of thehybrid derived by crossing hybrid rice HR120001 as a pollen donor. Thus,any methods using hybrid rice HR120001 in backcrosses, hybridproduction, crosses to populations, and the like, are part of thisinvention. All plants which are a progeny of or descend from hybrid riceHR120001 are within the scope of this invention. It is an aspect of thisinvention for hybrid rice HR120001 to be used in crosses with other,different, rice plants to produce first generation (F₁) rice hybridseeds and plants.

In another aspect, the present invention provides for single gene ormultiple gene converted plants of the parents of hybrid rice HR120001.The single or multiple transferred gene(s) may preferably be a dominantor recessive allele. Preferably, the single or multiple transferredgene(s) will confer such traits as herbicide resistance, insectresistance, resistance to bacterial, fungal, or viral diseases, malefertility, male sterility, enhanced nutritional quality, and industrialusage. The single or multiple gene(s) may be a naturally occurring ricegene or a transgene introduced through genetic engineering techniques.

In another aspect, the present invention provides regenerable cells foruse in tissue culture of hybrid rice plant HR120001. The tissue culturewill preferably be capable of regenerating plants having thephysiological and morphological characteristics of the foregoing riceplant, and of regenerating plants having substantially the same genotypeas the foregoing rice plant. Genetic variants of hybrid rice plantHR120001 naturally generated through using tissue culture orartificially induced utilizing mutagenic agents during tissue cultureare aspects of the present invention. Preferably, the regenerable cellsin such tissue cultures will be embryos, protoplasts, meristematiccells, callus, pollen, cotyledon, leaves, flowers, anthers, roots,pistils, root tips, glumes, seeds, panicles or stems. Still further, thepresent invention provides rice plants regenerated from the tissuecultures of the invention.

In another aspect, the present invention provides for producing a blendconsisting of rice seed of rice hybrid HR120001 with rice seed of riceinbred, rice variety or rice hybrid. The blend may also include a firstquantity of seed of rice hybrid HR120001 with one, two, three, four,five or more quantities of rice seed of another rice hybrid, rice inbredor rice variety.

In another aspect, the present invention also provides for producing ablend of seed of rice hybrid HR120001 with seed of one, two, three,four, five or more of another rice hybrid, rice variety or rice inbredwhere rice hybrid HR120001 is present in proportions from 1% up to 95%of the blend. Another aspect of this invention is planting the blendproduced with seeds of rice hybrid HR120001 and seeds of one, two three,four, five or more of another rice hybrid, rice variety or rice inbredand obtaining a crop with a mix of plants with rice hybrid HR120001 as acomponent. Further, another aspect of this invention is the harvest ofseeds from a planted blend of plants of which rice hybrid HR120001 is acomponent of the blend for the purpose of utilizing such seeds for food,feed, as a raw material in industry or as a seed source for planting.

In addition to the exemplary aspects and embodiments described above,further aspects and embodiments will become apparent by study of thefollowing descriptions.

DETAILED DESCRIPTION OF THE INVENTION

In the description and tables which follow, a number of terms are used.In order to provide a clear and consistent understanding of thespecification and claims, including the scope to be given such terms,the following definitions are provided:

Alkali Spreading Value. A 1-7 index used as predictor of starchgelatinization temperature and established by the extent ofdisintegration of milled rice kernel in contact with a dilute alkalisolution. Standard long grains have a 3 to 5 Alkali Spreading Value.

Allele. Allele is any one of many alternative forms of a gene, all ofwhich relate to one trait or characteristic. In a diploid cell ororganism, the two alleles of a given gene occupy corresponding loci on apair of homologous chromosomes.

Alter. The utilization of up-regulation, down-regulation, or genesilencing.

Amylose. Type of grain starch that affects cooking behavior. As such itsmeasured quantity in rice is used to establish cooking properties ofStandard US grain classes, or types. (long, medium and short grain).

Apparent Amylose Percent. The percentage of the endosperm starch ofmilled rice that is amylose. Standard long grains contain 20 to 23percent amylose. Rexmont-type long grains contain 24 to 25 percentamylose. Short and medium grains contain 14 to 16 percent amylose. Waxyrice contains zero percent amylose. Amylose values, like mostcharacteristics of rice, will vary over environments. “Apparent” refersto the procedure for determining amylose, which may also involvemeasuring some long chain amylopectin molecules that bind to some of theamylose molecules. These amylopectin molecules actually act similar toamylose in determining the relative hard or soft cookingcharacteristics.

Backcrossing. Process of crossing a hybrid progeny to one of theparents, for example, a first generation hybrid F₁ with one of theparental genotypes of the F₁ hybrid.

Blend. Physically mixing rice seeds of a rice hybrid with seeds of one,two, three, four or more of another rice hybrid, rice variety or riceinbred. A blend of rice seed can, for example, also include a mixture ofrice seed of rice hybrid HR120001 with rice seeds of one, two, three,four, five or more of another rice hybrid, rice variety or rice inbred.Planting a blend of rice seed is comprised of planting, for example,seeds of rice hybrid HR120001 with rice seeds of one, two, three, four,five or more of another rice hybrid, rice inbred or rice variety toproduce a crop containing the characteristics of all of the rice seedsand plants in this blend.

Breakdown. The Peak Viscosity minus the Trough Viscosity.

Cell. Cell as used herein includes a plant cell, whether isolated, intissue culture or incorporated in a plant or plant part.

Chalk. An opaque region of the rice kernel resulting from loose packingof the starch granules. Chalk may occur throughout or in a part of thekernel.

Consistency. The Final Viscosity minus the Trough Viscosity.

Cotyledon. A cotyledon is a type of seed leaf. The cotyledon containsthe food storage tissues of the seed.

Days to 50% heading. Number of days from emergence to the day when 50%of all panicles are exerted at least partially through the leaf sheath.A measure of growth duration.

Embryo. The embryo is the small plant contained within a mature seed.

Essentially all the physiological and morphological characteristics. Aplant having essentially all the physiological and morphologicalcharacteristics of the hybrid or cultivar, except for thecharacteristics derived from the converted gene.

Final Viscosity. The stickiness of rice flour/water slurry after beingheated to 95° C. and uniformly cooled to 50° C. in a standardizedinstrument, specifically the RAPID VISCO Analyzer. Viscosity at the endof the test also defined as Cool Paste Viscosity. (AACC Method 61-02)

Grain Length (L). Length of a whole rice grain measured in millimeters.

Gelatinization temperature. The temperature at which the consistency ofa rice flour-water mixture changes into a jelly. Correlates with thecooking time and texture of a rice product.

Gene Silencing. The interruption or suppression of the expression of agene at the level of transcription or translation.

Genetically Modified. Describes an organism that has received geneticmaterial from another, or had its genetic material modified, resultingin a change in one or more of its phenotypic characteristics. Methodsused to modify, introduce or delete the genetic material may includemutation breeding, backcross conversion, genetic transformation, singleand multiple gene conversion, and/or direct gene transfer.

Genotype. Refers to the genetic constitution of a cell or organism.

Grain Width (W). Width of a whole rice grain measured in millimeters.

Grain Yield. Weight of grain harvested from a given area. Grain yieldcould also be determined indirectly by multiplying the number ofpanicles per area, by the number of grains per panicle, and by grainweight.

Harvest Moisture. The percent of moisture of the grain when harvested.

Length/Width (L/W) Ratio. This ratio is determined by dividing theaverage length (L) by the average width (W).

Linkage. Refers to a phenomenon wherein alleles on the same chromosometend to segregate together more often than expected by chance if theirtransmission was independent.

Linkage Disequilibrium. Refers to a phenomenon wherein alleles tend toremain together in linkage groups when segregating from parents tooffspring, with a greater frequency than expected from their individualfrequencies.

Locus. A locus confers one or more traits such as, for example, malesterility, herbicide tolerance, insect resistance, disease resistance,waxy starch, modified fatty acid metabolism, modified phytic acidmetabolism, modified carbohydrate metabolism and modified proteinmetabolism. The trait may be, for example, conferred by a naturallyoccurring gene introduced into the genome of the variety bybackcrossing, a natural or induced mutation, or a transgene introducedthrough genetic transformation techniques. A locus may comprise one ormore alleles integrated at a single chromosomal location.

Lodging Percent. Lodging is a subjective measured rating, and is thepercentage of plant stems leaning or fallen completely to the groundbefore harvest.

Mixing. Physically mixing whole seeds of two or more genotypes of riceseed. For example, one of the genotypes of rice seed is rice hybridHR120001 mixed with another one, two, three, four, five or moregenotypes of rice seed.

Multiple Gene Converted (Conversion). Multiple gene converted(conversion) includes plants developed by a plant breeding techniquecalled backcrossing wherein essentially all of the desired morphologicaland physiological characteristics of an inbred are recovered, whileretaining two or more genes transferred into the inbred via crossing andbackcrossing. The term can also refer to the introduction of multiplegenes through genetic engineering techniques known in the art.

1000 Grain Wt. The weight of 1000 rice grains as measured in grams.

Paste Temperature. The temperature at which a defined flour-watermixture exhibits an initial viscosity increase under a standardizedprotocol utilizing the RAPID VISCO Analyzer. Paste Temperature is anindication of gelatinization temperature.

Paste Time. The time at which Paste Temperature occurs.

Peak Temperature. The temperature at which Peak Viscosity is attained.

Peak Time. The time at which Peak Viscosity is attained.

Peak Viscosity. The maximum viscosity attained during heating when astandardized protocol utilizing the RAPID VISCO Analyzer is applied to adefined rice flour-water slurry. (AACC Method 61-02).

Percent Identity. Percent identity as used herein refers to thecomparison of the homozygous alleles of two rice varieties. Percentidentity is determined by comparing a statistically significant numberof the homozygous alleles of two developed varieties. For example, apercent identity of 90% between rice variety 1 and rice variety 2 meansthat the two varieties have the same allele at 90% of their loci.

Percent Similarity. Percent similarity as used herein refers to thecomparison of the homozygous alleles of a rice variety with another riceplant, and if the homozygous allele of both rice plants matches at leastone of the alleles from the other plant then they are scored as similar.Percent similarity is determined by comparing a statisticallysignificant number of loci and recording the number of loci with similaralleles as a percentage. A percent similarity of 90% between the riceplant of this invention and another plant means that the rice plant ofthis invention matches at least one of the alleles of the other riceplant at 90% of the loci.

Plant. As used herein, the term “plant” includes reference to animmature or mature whole plant, including a plant from which seed orgrain or anthers have been removed. Seed or embryo that will produce theplant is also considered to be the plant.

Plant Height. Plant height in centimeters is taken from soil surface tothe tip of the extended panicle at harvest.

Plant Part. As used herein, the term “plant part” (or a rice plant, or apart thereof) includes protoplasts, leaves, stems, roots, root tips,anthers, seed, grain, embryo, pollen, ovules, cotyledon, hypocotyl,glumes, panicles, flower, shoot, tissue, cells, meristematic cells andthe like.

Pubescence. This refers to a covering of very fine hairs closelyarranged on the leaves, stems and glumes of the rice plant.

Quantitative Trait Loci (QTL). Genetic loci that controls to some degreenumerically measurable traits that are usually continuously distributed.

Regeneration. Regeneration refers to the development of a plant fromtissue culture.

RVA. RAPID VISCO Analyzer is a widely used laboratory instrumentutilized to examine the cooking properties of rice flour (i.e. pastetime and thickening ability).

RVU. RAPID VISCO units refers to the measurement units of the RVA.

Sakate Milling Degree meter. A milling meter that simultaneouslymeasures the degree of milling, comparative whiteness and degree oftransparency of milled rice samples.

Setback. The Final Viscosity minus Peak Viscosity.

Single Gene Converted (Conversion). Single gene converted (conversion)includes plants developed by a plant breeding technique calledbackcrossing wherein essentially all of the desired morphological andphysiological characteristics of an inbred are recovered, whileretaining a single gene transferred into the inbred via crossing andbackcrossing. The term can also refer to the introduction of a singlegene through genetic engineering techniques known in the art.

Total Milling (also call Milling Yield). The quantity of total milledrice produced in the milling of rough rice to a well-milled degree; itis usually expressed as a percent of rough rice by weight, but whenspecified, may be expressed as a percent of brown rice.

Trough Time. The time at which Trough Viscosity is attained.

Trough Viscosity. The minimum viscosity that occurs after Peak viscositywhen a standardized protocol utilizing the Rapid Visco Analyzer isapplied to a defined rice flour-water slurry. (AACC Method 61-02)

Whole Milling (also called Head Rice Milling Yield). The quantity ofmilled head (¾ to whole kernels) rice produced in the milling of roughrice to a well-milled degree, usually expressed in the United States asa percent of rough rice by weight.

Rice hybrid HR120001 is a high yielding, very early maturing, andphotoperiod insensitive long grain hybrid rice. The hybrid has shownuniformity and stability, as described in the following hybriddescription information. It has been produced and tested a sufficientnumber of years with careful attention to uniformity of plant type. Ricehybrid HR120001 has been produced with continued observation foruniformity of the parent lines.

Rice hybrid HR120001 has the following morphologic and othercharacteristics (based primarily on data collected at Alvin, Tex.).

TABLE 1 HYBRID DESCRIPTION INFORMATION Maturity (Alvin, Texas at 150kg/ha N): Days to maturity: 74 days from emergence to 50% headingMaturity Class: Very early (less than 75 days) Culm (Degrees fromperpendicular after flowering): Angle: Erect Length: 97.2 cm (Soil levelto top of extended panicle on main stem) Height Class: Short (90-100 cm)Internode Color (After flowering): Light gold Lodging Index: 0 - Strongresistance to lodging Flag Leaf (After Heading): Length: 35.7 cm Width:1.9 cm Pubescence: Pubescences present Leaf Angle (After heading):Intermediate Blade Color: Purple margins Basal Leaf Sheath Color: Lightpurple to purple Ligule: Length: 22.2 mm Color (Late vegetative state):Purple lines Shape: Cleft, tip is split Collar Color (Late vegetativestage): Green Auricle Color (Late vegetative stage): Pale green Panicle:Length: 24 cm Type: Intermediate Secondary Branching: Light Exertion(near maturity): Well, panicle base appears way above the collar of theflag leaf Axis: Droopy Shattering: Moderate Threshability: IntermediateGrain (Spikelet): Awns (After full heading): Absent Apiculus Color (Atmaturity): Straw Stigma Color: Purple Stigma Exertion (at flowering):100% exertion, stigma fully outside glume Lemma and Palea Color (At Goldand/or gold furrows on straw maturity): background Lemma and PaleaPubescence: Short hairs Spikelet Sterility (At maturity): Highly fertile(>90%) Grain (Seed): Seed Coat Color: Light brown Endosperm Type:Nonglutinous (nonwaxy) Endosperm Translucency: Clear EndospermChalkiness: Small (Less than 10% of grains with more than 50% ofchalkiness) Scent: Nonscented Shape Class (Length/width ratio): Longgrain Measurements (Milled): Length: 6.7 mm Width: 2.09 mm L/W ratio:3.21 Weight (1000 grains): 17 g Milling Yield (% whole kernel 66.3%(head) rice to rough rice): Apparent Amylose: 24% Alkali SpreadingValue: 3.67 (1.5% KOH Solution) Gelatinization Temperature Type:Intermediate Disease Resistance: Rice Blast (Pyricularia oryzae):Resistant (evaluations were conducted based on inoculations made with amix of the most predominant P. oryzae pathotypes found in the southernUS rice growing area) Sheath Blight (Rhizoctonia solani): ModeratelyResistant

This invention is also directed to methods for producing a rice plant bycrossing a first parent rice plant with a second parent rice plant,wherein the first or second rice plant is a rice plant from hybrid riceHR120001. Further, both first and second parent rice plants may be fromthe hybrid rice HR120001. Therefore, any methods using hybrid riceHR120001 are part of this invention: selfing, backcrosses, hybridbreeding, and crosses to populations. Any plants produced using hybridrice HR120001 as a parent is within the scope of this invention.

Still further, this invention also is directed to methods for producinga hybrid rice HR120001-derived rice plant by crossing rice hybridHR120001 with a second rice plant and growing the progeny seed, andrepeating the crossing and growing steps with rice hybridHR120001-derived plant from 0 to 7 times. Thus, any such methods usingthe rice hybrid HR120001 are part of this invention: selfing,backcrosses, hybrid production, crosses to populations, and the like.All plants produced using rice hybrid HR120001 as a parent are withinthe scope of this invention, including plants derived from rice hybridHR120001.

It should be understood that the parents of hybrid rice HR120001 can,through routine manipulation of cytoplasmic or other factors, beproduced in a male-sterile form. Such embodiments are also contemplatedwithin the scope of the present claims.

FURTHER EMBODIMENTS OF THE INVENTION

The advent of new molecular biological techniques has allowed theisolation and characterization of genetic elements with specificfunctions such as encoding specific protein products. Scientists in thefield of plant biology developed a strong interest in engineering thegenome of plants to contain and express foreign genetic elements, oradditional, or modified versions of native or endogenous in order toalter the traits of a plant in a specific manner. Any DNA sequenceswhether from a different species or from the same species which areinserted into the genome via transformation are referred to hereincollectively as “transgenes”. In some embodiments of the invention, atransgenic variant of rice hybrid HR120001 may contain at least onetransgene but could contain at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10and/or no more than 15, 14, 13, 12, 11, 10, 9, 8, 7, 6, 5, 4, 3, or 2.Over the last fifteen to twenty years, several methods for producingtransgenic plants have been developed, and the present invention, inparticular embodiments, also relates to transformed versions of theparents of the claimed hybrid.

Culture for expressing desired structural genes and cultured cells areknown in the art. Also as known in the art, rice is transformable andregenerable such that whole plants containing and expressing desiredgenes under regulatory control may be obtained. General descriptions ofplant expression vectors and reporter genes and transformation protocolscan be found in Gruber, et al., “Vectors for Plant Transformation”, inMethods in Plant Molecular Biology & Biotechnology, Glich, et al., (Eds.pp. 89-119, CRC Press, 1993). Moreover GUS expression vectors and GUSgene cassettes are available from Clone Tech Laboratories, Inc., PaloAlto, Calif. while luciferase expression vectors and luciferase genecassettes are available from ProMega Corp. (Madison, Wis.). Generalmethods of culturing plant tissues are provided for example by Maki, etal., “Procedures for Introducing Foreign DNA into Plants” in Methods inPlant Molecular Biology & Biotechnology, Glich, et al., (Eds. pp. 67-88CRC Press, 1993); and by Phillips, et al., “Cell-Tissue Culture andIn-Vitro Manipulation” in Corn & Corn Improvement, 3rd Edition; Sprague,et al., (Eds. pp. 345-387) American Society of Agronomy Inc., 1988.Methods of introducing expression vectors into plant tissue include thedirect infection or co-cultivation of plant cells with Agrobacteriumtumefaciens, Horsch et al., Science, 227:1229 (1985). Descriptions ofAgrobacterium vector systems and methods for Agrobacterium-mediated genetransfer provided by Gruber, et al., supra.

One embodiment of the invention is a process for producing rice hybridHR120001 further comprising a desired trait, said process comprisingtransforming a rice hybrid plant of HR120001 with a transgene thatconfers a desired trait. Another embodiment is the product produced bythis process. In one embodiment the desired trait may be one or more ofherbicide resistance, insect resistance, disease resistance, decreasedphytate, or modified fatty acid or carbohydrate metabolism. The specificgene may be any known in the art or listed herein, including; apolynucleotide conferring resistance to imidazolinone, sulfonylurea,glyphosate, glufosinate, triazine, benzonitrile, cyclohexanedione,phenoxy proprionic acid and L-phosphinothricin; a polynucleotideencoding a Bacillus thuringiensis polypeptide, a polynucleotide encodingphytase, FAD-2, FAD-3, galactinol synthase or a raffinose syntheticenzyme.

Numerous methods for plant transformation have been developed, includingbiological and physical plant transformation protocols. See, forexample, Miki et al., “Procedures for Introducing Foreign DNA intoPlants” in Methods in Plant Molecular Biology and Biotechnology, Glick,B. R. and Thompson, J. E. Eds. (CRC Press, Inc., Boca Raton, 1993) pages67-88 and Armstrong, “The First Decade of Maize Transformation: A Reviewand Future Perspective” (Maydica 44:101-109, 1999). In addition,expression vectors and in vitro culture methods for plant cell or tissuetransformation and regeneration of plants are available. See, forexample, Gruber et al., “Vectors for Plant Transformation” in Methods inPlant Molecular Biology and Biotechnology, Glick, B. R. and Thompson, J.E. Eds. (CRC Press, Inc., Boca Raton, 1993) pages 89-119.

A genetic trait which has been engineered into the genome of aparticular rice plant may then be moved into the genome of another riceplant using traditional breeding techniques that are well known in theplant breeding arts. For example, a backcrossing approach is commonlyused to move a transgene from a transformed rice plant into an alreadydeveloped rice hybrid or variety, and the resulting backcross conversionplant would then comprise the transgene(s).

Plant transformation involves the construction of an expression vectorwhich will function in plant cells. Such a vector comprises DNAcomprising a gene under control of or operatively linked to a regulatoryelement (for example, a promoter). The expression vector may contain oneor more such operably linked gene/regulatory element combinations. Thevector(s) may be in the form of a plasmid, and can be used alone or incombination with other plasmids, to provide transformed rice plants,using transformation methods as described below to incorporatetransgenes into the genetic material of the rice plant(s).

Expression Vectors for Rice Transformation—Marker Genes

Expression vectors include at least one genetic marker, operably linkedto a regulatory element (a promoter, for example) that allowstransformed cells containing the marker to be either recovered bynegative selection, i.e., inhibiting growth of cells that do not containthe selectable marker gene, or by positive selection, i.e., screeningfor the product encoded by the genetic marker. Many commonly usedselectable marker genes for plant transformation are well known in thetransformation arts, and include, for example, genes that code forenzymes that metabolically detoxify a selective chemical agent which maybe an antibiotic or an herbicide, or genes that encode an altered targetwhich is insensitive to the inhibitor. A few positive selection methodsare also known in the art.

One commonly used selectable marker gene for plant transformation is theneomycin phosphotransferase II (nptII) gene, isolated from transposonTn5, which when placed under the control of plant regulatory signalsconfers resistance to kanamycin. Fraley et al., Proc. Natl. Acad. Sci.U.S.A., 80:4803 (1983). Another commonly used selectable marker gene isthe hygromycin phosphotransferase gene which confers resistance to theantibiotic hygromycin. Vanden Elzen et al., Plant Mol. Biol., 5:299(1985).

Additional selectable marker genes of bacterial origin that conferresistance to antibiotics include gentamycin acetyl transferase,streptomycin phosphotransferase, and aminoglycoside-3′-adenyltransferase, the bleomycin resistance determinant. Hayford et al., PlantPhysiol. 86:1216 (1988); Jones et al., Mol. Gen. Genet., 210:86 (1987);Svab et al., Plant Mol. Biol. 14:197 (1990); Hille et al., Plant Mol.Biol. 7:171 (1986). Other selectable marker genes confer resistance toherbicides such as glyphosate, glufosinate or bromoxynil. Comai et al.,Nature 317:741-744 (1985), Gordon-Kamm et al., Plant Cell 2:603-618(1990) and Stalker et al., Science 242:419-423 (1988).

Other selectable marker genes for plant transformation not of bacterialorigin include, for example, mouse dihydrofolate reductase, plant5-enolpyruvylshikimate-3-phosphate synthase and plant acetolactatesynthase. Eichholtz et al., Somatic Cell Mol. Genet. 13:67 (1987), Shahet al., Science 233:478 (1986), Charest et al., Plant Cell Rep. 8:643(1990).

Another class of marker genes for plant transformation requiresscreening of presumptively transformed plant cells rather than directgenetic selection of transformed cells for resistance to a toxicsubstance such as an antibiotic. These genes are particularly useful toquantify or visualize the spatial pattern of expression of a gene inspecific tissues and are frequently referred to as reporter genesbecause they can be fused to a gene or gene regulatory sequence for theinvestigation of gene expression. Commonly used genes for screeningpresumptively transformed cells include β-glucuronidase (GUS),β-galactosidase, luciferase and chloramphenicol acetyltransferase.Jefferson, R. A., Plant Mol. Biol. Rep. 5:387 (1987), Teeri et al., EMBOJ. 8:343 (1989), Koncz et al., Proc. Natl. Acad. Sci. U.S.A. 84:131(1987), DeBlock et al., EMBO J. 3:1681 (1984). Another approach to theidentification of relatively rare transformation events has been use ofa gene that encodes a dominant constitutive regulator of the Zea maysanthocyanin pigmentation pathway. Ludwig et al., Science 247:449 (1990).

In vivo methods for visualizing GUS activity that do not requiredestruction of plant tissue are available. Molecular Probes publication2908, IMAGENE GREEN, p. 1-4 (1993) and Naleway et al., J. Cell Biol.115:151a (1991). However, these in vivo methods for visualizing GUSactivity have not proven useful for recovery of transformed cellsbecause of low sensitivity, high fluorescent backgrounds and limitationsassociated with the use of luciferase genes as selectable markers.

More recently, a gene encoding Green Fluorescent Protein (GFP) has beenutilized as a marker for gene expression in prokaryotic and eukaryoticcells. Chalfie et al., Science 263:802 (1994). GFP and mutants of GFPmay be used as screenable markers.

Expression Vectors for Rice Transformation—Promoters

Genes included in expression vectors must be driven by nucleotidesequence comprising a regulatory element, for example, a promoter.Several types of promoters are now well known in the transformationarts, as are other regulatory elements that can be used alone or incombination with promoters.

As used herein, “promoter” includes reference to a region of DNAupstream from the start of transcription and involved in recognition andbinding of RNA polymerase and other proteins to initiate transcription.A “plant promoter” is a promoter capable of initiating transcription inplant cells. Examples of promoters under developmental control includepromoters that preferentially initiate transcription in certain tissues,such as leaves, roots, seeds, fibers, xylem vessels, tracheids, orsclerenchyma. Such promoters are referred to as “tissue-preferred”.Promoters which initiate transcription only in certain tissue arereferred to as “tissue-specific”. A “cell type” specific promoterprimarily drives expression in certain cell types in one or more organs,for example, vascular cells in roots or leaves. An “inducible” promoteris a promoter which is under environmental control. Examples ofenvironmental conditions that may effect transcription by induciblepromoters include anaerobic conditions or the presence of light.Tissue-specific, tissue-preferred, cell type specific, and induciblepromoters constitute the class of “non-constitutive” promoters. A“constitutive” promoter is a promoter which is active under mostenvironmental conditions.

A. Inducible Promoters—An inducible promoter is operably linked to agene for expression in rice. Optionally, the inducible promoter isoperably linked to a nucleotide sequence encoding a signal sequencewhich is operably linked to a gene for expression in rice. With aninducible promoter the rate of transcription increases in response to aninducing agent.

Any inducible promoter can be used in the instant invention. See Ward etal., Plant Mol. Biol. 22:361-366 (1993). Exemplary inducible promotersinclude, but are not limited to, that from the ACEI system whichresponds to copper (Meft et al., PNAS 90:4567-4571 (1993)); In2 genefrom maize which responds to benzenesulfonamide herbicide safeners(Hershey et al., Mol. Gen Genetics 227:229-237 (1991) and Gatz et al.,Mol. Gen. Genetics 243:32-38 (1994)) or Tet repressor from Tn10 (Gatz etal., Mol. Gen. Genetics 227:229-237 (1991)). A particularly preferredinducible promoter is a promoter that responds to an inducing agent towhich plants do not normally respond. An exemplary inducible promoter isthe inducible promoter from a steroid hormone gene, the transcriptionalactivity of which is induced by a glucocorticosteroid hormone (Schena etal., Proc. Natl. Acad. Sci. U.S.A. 88:0421 (1991)).

B. Constitutive Promoters—A constitutive promoter is operably linked toa gene for expression in rice or the constitutive promoter is operablylinked to a nucleotide sequence encoding a signal sequence which isoperably linked to a gene for expression in rice.

Many different constitutive promoters can be utilized in the instantinvention. Exemplary constitutive promoters include, but are not limitedto, the promoters from plant viruses such as the 35S promoter from CaMV(Odell et al., Nature 313:810-812 (1985)) and the promoters from suchgenes as rice actin (McElroy et al., Plant Cell 2:163-171 (1990));ubiquitin (Christensen et al., Plant Mol. Biol. 12:619-632 (1989) andChristensen et al., Plant Mol. Biol. 18:675-689 (1992)); pEMU (Last etal., Theor. Appl. Genet. 81:581-588 (1991)); MAS (Velten et al., EMBO J.3:2723-2730 (1984)) and maize H3 histone (Lepetit et al., Mol. Gen.Genetics 231:276-285 (1992) and Atanassova et al., Plant Journal 2 (3):291-300 (1992)).

The ALS promoter, Xba1/Nco1 fragment 5′ to the Brassica napus ALS3structural gene (or a nucleotide sequence similarity to said Xba1/Nco1fragment), represents a particularly useful constitutive promoter. SeePCT application WO 96/30530.

C. Tissue-specific or Tissue-preferred Promoters—A tissue-specificpromoter is operably linked to a gene for expression in rice.Optionally, the tissue-specific promoter is operably linked to anucleotide sequence encoding a signal sequence which is operably linkedto a gene for expression in rice. Plants transformed with a gene ofinterest operably linked to a tissue-specific promoter produce theprotein product of the transgene exclusively, or preferentially, in aspecific tissue.

Any tissue-specific or tissue-preferred promoter can be utilized in theinstant invention. Exemplary tissue-specific or tissue-preferredpromoters include, but are not limited to, a root-preferred promoter,such as that from the phaseolin gene (Murai et al., Science 23:476-482(1983) and Sengupta-Gopalan et al., Proc. Natl. Acad. Sci. U.S.A.82:3320-3324 (1985)); a leaf-specific and light-induced promoter such asthat from cab or rubisco (Simpson et al., EMBO J. 4(11):2723-2729 (1985)and Timko et al., Nature 318:579-582 (1985)); an anther-specificpromoter such as that from LAT52 (Twell et al., Mol. Gen. Genetics217:240-245 (1989)); a pollen-specific promoter such as that from Zm13(Guerrero et al., Mol. Gen. Genetics 244:161-168 (1993)) or amicrospore-preferred promoter such as that from apg (Twell et al., Sex.Plant Reprod. 6:217-224 (1993)).

Signal Sequences for Targeting Proteins to Subcellular Compartments

Transport of a protein produced by transgenes to a subcellularcompartment such as the chloroplast, vacuole, peroxisome, glyoxysome,cell wall or mitochondrion or for secretion into the apoplast, isaccomplished by means of operably linking the nucleotide sequenceencoding a signal sequence to the 5′ and/or 3′ region of a gene encodingthe protein of interest. Targeting sequences at the 5′ and/or 3′ end ofthe structural gene may determine, during protein synthesis andprocessing, where the encoded protein is ultimately compartmentalized.

The presence of a signal sequence directs a polypeptide to either anintracellular organelle or subcellular compartment or for secretion tothe apoplast. Many signal sequences are known in the art. See, forexample Becker et al., Plant Mol. Biol. 20:49 (1992); Knox, C., et al.,Plant Mol. Biol. 9:3-17 (1987); Lerner et al., Plant Physiol. 91:124-129(1989); Fontes et al., Plant Cell 3:483-496 (1991); Matsuoka et al.,Proc. Natl. Acad. Sci. 88:834 (1991); Gould et al., J. Cell. Biol.108:1657 (1989); Creissen et al., Plant J. 2:129 (1991); Kalderon, etal., Cell 39:499-509 (1984); Steifel, et al., Plant Cell 2:785-793(1990).

Foreign Protein Genes and Agronomic Genes

With transgenic plants according to the present invention, a foreignprotein can be produced in commercial quantities. Thus, techniques forthe selection and propagation of transformed plants, which are wellunderstood in the art, yield a plurality of transgenic plants which areharvested in a conventional manner, and a foreign protein then can beextracted from a tissue of interest or from total biomass. Proteinextraction from plant biomass can be accomplished by known methods whichare discussed, for example, by Heney and Orr, Anal. Biochem. 114:92-6(1981).

According to a preferred embodiment, the transgenic plant provided forcommercial production of foreign protein is rice. In another preferredembodiment, the biomass of interest is seed. For the relatively smallnumber of transgenic plants that show higher levels of expression, agenetic map can be generated, primarily via conventional RFLP, PCR andSSR analysis, which identifies the approximate chromosomal location ofthe integrated DNA molecule. For exemplary methodologies in this regard,see Glick and Thompson in Methods in Plant Molecular Biology andBiotechnology CRC Press, Boca Raton 269:284 (1993).

Map information concerning chromosomal location is useful forproprietary protection of a subject transgenic plant. If unauthorizedpropagation is undertaken and crosses made with other germplasm, the mapof the integration region can be compared to similar maps for suspectplants, to determine if the latter have a common parentage with thesubject plant. Map comparisons would involve hybridizations, RFLP, PCR,SSR and sequencing, all of which are conventional techniques.

Through the transformation of rice, the expression of genes can bealtered to enhance disease resistance, insect resistance, herbicideresistance, agronomic quality and other traits. Transformation can alsobe used to insert DNA sequences which control or help controlmale-sterility. DNA sequences native to rice as well as non-native DNAsequences can be transformed into rice and used to alter levels ofnative or non-native proteins. Various promoters, targeting sequences,enhancing sequences, and other DNA sequences can be inserted into thegenome for the purpose of altering the expression of proteins. Reductionof the activity of specific genes (also known as gene silencing, or genesuppression) is desirable for several aspects of genetic engineering inplants.

Many techniques for gene silencing are well known to one of skill in theart, including but not limited to knock-outs (such as by insertion of atransposable element such as mu (Vicki Chandler, The Maize Handbook ch.118 (Springer-Verlag 1994) or other genetic elements such as a FRT, Loxor other site specific integration site, antisense technology (see,e.g., Sheehy et al. (1988) PNAS USA 85:8805-8809; and U.S. Pat. Nos.5,107,065; 5,453,566; and 5,759,829); co-suppression (e.g., Taylor(1997) Plant Cell 9:1245; Jorgensen (1990) Trends Biotech.8(12):340-344; Flavell (1994) PNAS USA 91:3490-3496; Finnegan et al.(1994) Bio/Technology 12: 883-888; and Neuhuber et al. (1994) Mol. Gen.Genet. 244:230-241); RNA interference (Napoli et al. (1990) Plant Cell2:279-289; U.S. Pat. No. 5,034,323; Sharp (1999) Genes Dev. 13:139-141;Zamore et al. (2000) Cell 101:25-33; and Montgomery et al. (1998) PNASUSA 95:15502-15507), virus-induced gene silencing (Burton, et al. (2000)Plant Cell 12:691-705; and Baulcombe (1999) Curr. Op. Plant Bio.2:109-113); target-RNA-specific ribozymes (Haseloff et al. (1988) Nature334: 585-591); hairpin structures (Smith et al. (2000) Nature407:319-320; WO 99/53050; and WO 98/53083); MicroRNA (Aukerman & Sakai(2003) Plant Cell 15:2730-2741); ribozymes (Steinecke et al. (1992) EMBOJ. 11:1525; and Perriman et al. (1993) Antisense Res. Dev. 3:253);oligonucleotide mediated targeted modification (e.g., WO 03/076574 andWO 99/25853); Zn-finger targeted molecules (e.g., WO 01/52620; WO03/048345; and WO 00/42219); and other methods or combinations of theabove methods known to those of skill in the art.

Likewise, by means of the present invention, agronomic genes can beexpressed in transformed plants. More particularly, plants can begenetically engineered to express various phenotypes of agronomicinterest. Exemplary genes implicated in this regard include, but are notlimited to, those categorized below:

1. Genes that Confer Resistance to Pests or Disease and that Encode:

A. Plant disease resistance genes. Plant defenses are often activated byspecific interaction between the product of a disease resistance gene(R) in the plant and the product of a corresponding avirulence (Avr)gene in the pathogen. A plant cultivar can be transformed with clonedresistance gene to engineer plants that are resistant to specificpathogen strains. See, for example Jones et al., Science 266:789 (1994)(cloning of the tomato Cf-9 gene for resistance to Cladosporium falvum);Martin et al., Science 262:1432 (1993) (tomato Pto gene for resistanceto Pseudomonas syringae pv. tomato encodes a protein kinase); Mindrinoset al., Cell 78:1089 (1994) (Arabidopsis RSP2 gene for resistance toPseudomonas syringae); McDowell & Woffenden, (2003) Trends Biotechnol.21(4): 178-83 and Toyoda et al., (2002) Transgenic Res. 11 (6):567-82.

B. A Bacillus thuringiensis protein, a derivative thereof or a syntheticpolypeptide modeled thereon. See, for example, Geiser et al., Gene48:109 (1986), who disclose the cloning and nucleotide sequence of a Btδ-endotoxin gene. Moreover, DNA molecules encoding δ-endotoxin genes canbe purchased from American Type Culture Collection, Manassas, Va., forexample, under ATCC Accession Nos. 40098, 67136, 31995 and 31998.

C. A lectin. See, for example, the disclosure by Van Damme et al., PlantMolec. Biol. 24:25 (1994), who disclose the nucleotide sequences ofseveral Clivia miniata mannose-binding lectin genes.

D. A vitamin-binding protein such as avidin. See PCT application US93/06487. The application teaches the use of avidin and avidinhomologues as larvicides against insect pests.

E. An enzyme inhibitor, for example, a protease or proteinase inhibitoror an amylase inhibitor. See, for example, Abe et al., J. Biol. Chem.262:16793 (1987) (nucleotide sequence of rice cysteine proteinaseinhibitor), Huub et al., Plant Molec. Biol. 21:985 (1993) (nucleotidesequence of cDNA encoding tobacco proteinase inhibitor I), Sumitani etal., Biosci. Biotech. Biochem. 57:1243 (1993) (nucleotide sequence ofStreptomyces nitrosporeus α-amylase inhibitor).

F. An insect-specific hormone or pheromone such as an ecdysteroid andjuvenile hormone, a variant thereof, a mimetic based thereon, or anantagonist or agonist thereof. See, for example, the disclosure byHammock et al., Nature 344:458 (1990), of baculovirus expression ofcloned juvenile hormone esterase, an inactivator of juvenile hormone.

G. An insect-specific peptide or neuropeptide which, upon expression,disrupts the physiology of the affected pest. For example, see thedisclosures of Regan, J. Biol. Chem. 269:9 (1994) (expression cloningyields DNA coding for insect diuretic hormone receptor), and Pratt etal., Biochem. Biophys. Res. Comm. 163:1243 (1989) (an allostatin isidentified in Diploptera puntata). Chattopadhyay et al. (2004) CriticalReviews in Microbiology 30 (1): 33-54 2004; Zjawiony (2004) J Nat Prod67 (2): 300-310; Carlini & Grossi-de-Sa (2002) Toxicon, 40 (11):1515-1539; Ussuf et al. (2001) Curr Sci. 80 (7): 847-853; andVasconcelos & Oliveira (2004) Toxicon 44 (4): 385-403. See also U.S.Pat. No. 5,266,317 to Tomalski et al., who disclose genes encodinginsect-specific, paralytic neurotoxins.

H. An insect-specific venom produced in nature by a snake, a wasp, etc.For example, see Pang et al., Gene 116:165 (1992), for disclosure ofheterologous expression in plants of a gene coding for a scorpioninsectotoxic peptide.

I. An enzyme responsible for a hyper-accumulation of a monoterpene, asesquiterpene, a steroid, a hydroxamic acid, a phenylpropanoidderivative or another non-protein molecule with insecticidal activity.

J. An enzyme involved in the modification, including thepost-translational modification, of a biologically active molecule; forexample, a glycolytic enzyme, a proteolytic enzyme, a lipolytic enzyme,a nuclease, a cyclase, a transaminase, an esterase, a hydrolase, aphosphatase, a kinase, a phosphorylase, a polymerase, an elastase, achitinase and a glucanase, whether natural or synthetic. See PCTapplication WO 93/02197 in the name of Scott et al., which discloses thenucleotide sequence of a callase gene. DNA molecules which containchitinase-encoding sequences can be obtained, for example, from the ATCCunder Accession Nos. 39637 and 67152. See also Kramer et al., InsectBiochem. Molec. Biol. 23:691 (1993), who teach the nucleotide sequenceof a cDNA encoding tobacco hornworm chitinase, and Kawalleck et al.,Plant Molec. Biol. 21:673 (1993), who provide the nucleotide sequence ofthe parsley ubi4-2 polyubiquitin gene, U.S. Pat. Nos. 7,145,060,7,087,810 and 6,563,020

K. A molecule that stimulates signal transduction. For example, see thedisclosure by Botella et al., Plant Molec. Biol. 24:757 (1994), ofnucleotide sequences for mung bean calmodulin cDNA clones, and Griess etal., Plant Physiol. 104:1467 (1994), who provide the nucleotide sequenceof a maize calmodulin cDNA clone.

L. A hydrophobic moment peptide. See PCT application WO 95/16776 andU.S. Pat. No. 5,580,852 which discloses peptide derivatives ofTachyplesin which inhibit fungal plant pathogens and PCT application WO95/18855 and U.S. Pat. No. 5,607,914 which teaches syntheticantimicrobial peptides that confer disease resistance, the respectivecontents of which are hereby incorporated by reference.

M. A membrane permease, a channel former or a channel blocker. Forexample, see the disclosure of Jaynes et al., Plant Sci 89:43 (1993), ofheterologous expression of a cecropin-β, lytic peptide analog to rendertransgenic tobacco plants resistant to Pseudomonas solanacearum.

N. A viral-invasive protein or a complex toxin derived therefrom. Forexample, the accumulation of viral coat proteins in transformed plantcells imparts resistance to viral infection and/or disease developmenteffected by the virus from which the coat protein gene is derived, aswell as by related viruses. See Beachy et al., Ann. Rev. Phytopathol.28:451 (1990). Coat protein-mediated resistance has been conferred upontransformed plants against alfalfa mosaic virus, cucumber mosaic virus,tobacco streak virus, potato virus X, potato virus Y, tobacco etchvirus, tobacco rattle virus and tobacco mosaic virus. Id.

O. An insect-specific antibody or an immunotoxin derived therefrom.Thus, an antibody targeted to a critical metabolic function in theinsect gut would inactivate an affected enzyme, killing the insect. Cf.Taylor et al., Abstract #497, Seventh Int'l Symposium on MolecularPlant-Microbe Interactions (Edinburgh, Scotland) (1994) (enzymaticinactivation in transgenic tobacco via production of single-chainantibody fragments).

P. A virus-specific antibody. See, for example, Tavladoraki et al.,Nature 366:469 (1993), who show that transgenic plants expressingrecombinant antibody genes are protected from virus attack.

Q. A developmental-arrestive protein produced in nature by a pathogen ora parasite. Thus, fungal endo-α-1,4-D-polygalacturonases facilitatefungal colonization and plant nutrient release by solubilizing plantcell wall homo-α-1,4-D-galacturonase. See Lamb et al., Bio/Technology10:1436 (1992). The cloning and characterization of a gene which encodesa bean endopolygalacturonase-inhibiting protein is described by Toubartet al., Plant J. 2:367 (1992).

R. A developmental-arrestive protein produced in nature by a plant. Forexample, Logemann et al., Bio/Technology 10:305 (1992), have shown thattransgenic plants expressing the barley ribosome-inactivating gene havean increased resistance to fungal disease.

S. Genes involved in the Systemic Acquired Resistance (SAR) Responseand/or the pathogenesis-related genes. Briggs, S., Current Biology, 5(2)(1995); Pieterse & Van Loon (2004) Curr. Opin. Plant Bio. 7(4):456-64and Somssich (2003) Cell 113(7):815-6.

T. Antifungal genes. See Cornelissen and Melchers, Plant Physiol.,101:709-712 (1993); Parijs et al., Planta 183:258-264 (1991) andBushnell et al., Can. J. of Plant Path. 20(2):137-149 (1998). Also seeU.S. Pat. No. 6,875,907.

U. Detoxification genes, such as for fumonisin, beauvericin,moniliformin and zearalenone and their structurally related derivatives.For example, see U.S. Pat. No. 5,792,931.

V. Cystatin and cysteine proteinase inhibitors. See U.S. Pat. No.7,205,453.

W. Defensin genes. See WO 03/000863 and U.S. Pat. No. 6,911,577.

2. Genes that Confer Resistance to an Herbicide, for Example:

A. An herbicide that inhibits the growing point or meristem, such as animidazolinone or a sulfonylurea. Exemplary genes in this category codefor mutant ALS and AHAS enzyme as described, for example, by Lee et al.,EMBO J. 7:1241 (1988), and Miki et al., Theor. Appl. Genet. 80:449(1990), respectively.

B. Glyphosate (resistance conferred by mutant5-enolpyruvlshikimate-3-phosphate synthase (EPSP) and aroA genes,respectively and other phosphono compounds such as glufosinate(phosphinothricin acetyl transferase (PAT) and Streptomyceshygroscopicus PAT bar genes), and pyridinoxy or phenoxy propionic acidsand cyclohexanediones (ACCase inhibitor-encoding genes). See, forexample, U.S. Pat. No. 4,940,835 to Shah, et al., which discloses thenucleotide sequence of a form of EPSP which can confer glyphosateresistance. U.S. Pat. No. 5,627,061 to Barry et al. also describes genesencoding EPSPS enzymes. See also U.S. Pat. Nos. 6,566,587; 6,338,961;6,248,876 B1; 6,040,497; 5,804,425; 5,633,435; 5,145,783; 4,971,908;5,312,910; 5,188,642; 4,940,835; 5,866,775; 6,225,114 B1; 6,130,366;5,310,667; 4,535,060; 4,769,061; 5,633,448; 5,510,471; Re. 36,449; RE37,287 E; and 5,491,288; and international publications EP1173580; WO01/66704; EP1173581 and EP1173582, which are incorporated herein byreference for this purpose. Glyphosate resistance is also imparted toplants that express a gene that encodes a glyphosate oxido-reductaseenzyme as described more fully in U.S. Pat. Nos. 5,776,760 and5,463,175, which are incorporated herein by reference for this purpose.In addition glyphosate resistance can be imparted to plants by the overexpression of genes encoding glyphosate N-acetyltransferase. See, forexample, U.S. application Ser. No. 10/427,692. A DNA molecule encoding amutant aroA gene can be obtained under ATCC accession number 39256, andthe nucleotide sequence of the mutant gene is disclosed in U.S. Pat. No.4,769,061 to Comai. European patent application No. 0 333 033 to Kumadaet al., and U.S. Pat. No. 4,975,374 to Goodman et al., disclosenucleotide sequences of glutamine synthetase genes which conferresistance to herbicides such as L-phosphinothricin. The nucleotidesequence of a PAT gene is provided in European application No. 0 242 246to Leemans et al. DeGreef et al., Bio/Technology 7:61 (1989), describethe production of transgenic plants that express chimeric bar genescoding for PAT activity. Exemplary of genes conferring resistance tophenoxy propionic acids and cyclohexanedione, such as sethoxydim andhaloxyfop are the Acc1-S1, Acc1-S2 and Acc1-S3 genes described byMarshall et al., Theor. Appl. Genet. 83:435 (1992).

C. An herbicide that inhibits photosynthesis, such as a triazine (psbAand gs+ genes) or a benzonitrile (nitrilase gene). Przibilla et al.,Plant Cell 3:169 (1991), describe the transformation of Chlamydomonaswith plasmids encoding mutant psbA genes. Nucleotide sequences fornitrilase genes are disclosed in U.S. Pat. No. 4,810,648 to Stalker, andDNA molecules containing these genes are available under ATCC AccessionNos. 53435, 67441, and 67442. Cloning and expression of DNA coding for aglutathione S-transferase is described by Hayes et al., Biochem. J.285:173 (1992).

3. Genes that Confer or Contribute to a Value-Added Trait, Such as:

A. Modified fatty acid metabolism, for example, by transforming a plantwith an antisense gene of stearyl-ACP desaturase to increase stearicacid content of the plant. See Knultzon et al., Proc. Natl. Acad. Sci.U.S.A. 89:2624 (1992).

B. Decreased phytate content, 1) Introduction of a phytase-encoding genewould enhance breakdown of phytate, adding more free phosphate to thetransformed plant. For example, see Van Hartingsveldt et al., Gene127:87 (1993), for a disclosure of the nucleotide sequence of anAspergillus niger phytase gene; 2) Up-regulation of a gene that reducesphytate content. In maize, this, for example, could be accomplished bycloning and then re-introducing DNA associated with one or more of thealleles, such as the LPA alleles identified in maize mutantscharacterized by low levels of phytic acid. See Raboy et al., Maydica35:383 (1990) and/or by altering inositol kinase activity as ininternational publication numbers WO 02/059324, WO 03/027243, WO99/05298, WO 2002/059324, WO 98/45448, WO 99/55882, WO 01/04147; U.S.Publication Numbers 2003/0009011, 2003/0079247; and U.S. Pat. Nos.6,197,561, 6,291,224, 6,391,348.

C. Modified carbohydrate composition effected, for example, bytransforming plants with a gene coding for an enzyme that alters thebranching pattern of starch or a gene altering thioredoxin such as NTRand/or TRX (See U.S. Pat. No. 6,531,648 which is incorporated byreference for this purpose) and/or a gamma zein knock out or mutant suchas cs27 or TUSC27 or en27 (See U.S. Pat. No. 6,858,778 and U.S.Publication Nos. 2005/0160488 and 2005/0204418, which are incorporatedby reference for this purpose). See Shiroza et al., J. Bacteol. 170:810(1988) (nucleotide sequence of Streptococcus mutannsfructosyltransferase gene), Steinmetz et al., Mol. Gen. Genet. 20:220(1985) (nucleotide sequence of Bacillus subtilis levansucrase gene), Penet al., Bio/Technology 10:292 (1992) (production of transgenic plantsthat express Bacillus licheniformis α-amylase), Elliot et al., PlantMolec. Biol. 21:515 (1993) (nucleotide sequences of tomato invertasegenes), SØgaard et al., J. Biol. Chem. 268:22480 (1993) (site-directedmutagenesis of barley α-amylase gene), and Fisher et al., Plant Physiol.102:1045 (1993) (maize endosperm starch branching enzyme II), WO99/10498 (improved digestibility and/or starch extraction throughmodification of UDP-D-xylose 4-epimerase, Fragile 1 and 2, Ref 1, HCHL,C4H), U.S. Pat. No. 6,232,529 (method of producing high oil seed bymodification of starch levels (AGP)). The fatty acid modification genesmentioned above may also be used to affect starch content and/orcomposition through the interrelationship of the starch and oilpathways.

D. Altering conjugated linolenic or linoleic acid content, such as ininternational publication number WO 01/12800. Altering LEC1, AGP, Dek1,Superal1, mi1ps, various Ipa genes such as Ipa1, Ipa3, hpt or hggt. Forexample, see international publication numbers WO 02/42424, WO 98/22604,WO 03/011015, WO 02/057439, WO 03/011015; U.S. Pat. Nos. 6,423,886,6,197,561, 6,825,397, 7,157,621; U.S. Publication No. 2003/0079247 andRivera-Madrid, R. et al. Proc. Natl. Acad. Sci. 92:5620-5624 (1995).

E. Altered antioxidant content or composition, such as alteration oftocopherol or tocotrienols. For example, see U.S. Pat. Nos. 6,787,683and 7,154,029 and international publication number WO 00/68393 involvingthe manipulation of antioxidant levels through alteration of a phyt1prenyl transferase (ppt) and international publication number WO03/082899 through alteration of a homogentisate geranyl geranyltransferase (hggt).

F. Altered essential seed amino acids. For example, see U.S. Pat. No.6,127,600 (method of increasing accumulation of essential amino acids inseeds), U.S. Pat. No. 6,080,913 (binary methods of increasingaccumulation of essential amino acids in seeds), U.S. Pat. No. 5,990,389(high lysine), international publication number WO 99/40209 (alterationof amino acid compositions in seeds), international publication numberWO 99/29882 (methods for altering amino acid content of proteins), U.S.Pat. No. 5,850,016 (alteration of amino acid compositions in seeds),international publication number WO 98/20133 (proteins with enhancedlevels of essential amino acids), U.S. Pat. No. 5,885,802 (highmethionine), U.S. Pat. No. 5,885,801 (high threonine), U.S. Pat. No.6,664,445 (plant amino acid biosynthetic enzymes), U.S. Pat. No.6,459,019 (increased lysine and threonine), U.S. Pat. No. 6,441,274(plant tryptophan synthase beta subunit), U.S. Pat. No. 6,346,403(methionine metabolic enzymes), U.S. Pat. No. 5,939,599 (high sulfur),U.S. Pat. No. 5,912,414 (increased methionine), internationalpublication number WO 98/56935 (plant amino acid biosynthetic enzymes),international publication number WO 98/45458 (engineered seed proteinhaving higher percentage of essential amino acids), internationalpublication number WO 98/42831 (increased lysine), U.S. Pat. No.5,633,436 (increasing sulfur amino acid content), U.S. Pat. No.5,559,223 (synthetic storage proteins with defined structure containingprogrammable levels of essential amino acids for improvement of thenutritional value of plants), international publication number WO96/01905 (increased threonine), international publication number WO95/15392 (increased lysine), U.S. Pat. Nos. 6,930,225, 7,179,955,6,803,498, U.S. Publication No. 2004/0068767, international publicationnumbers WO 01/79516 and WO 00/09706 (Ces A: cellulose synthase), U.S.Pat. No. 6,194,638 (hemicellulose), U.S. Pat. Nos. 6,399,859 and7,098,381 (UDPGdH) and U.S. Pat. No. 6,194,638 (RGP).

4. Genes that Control Male Sterility

There are several methods of conferring genetic male sterilityavailable, such as multiple mutant genes at separate locations withinthe genome that confer male sterility, as disclosed in U.S. Pat. Nos.4,654,465 and 4,727,219 to Brar et al. and chromosomal translocations asdescribed by Patterson in U.S. Pat. Nos. 3,861,709 and 3,710,511. Inaddition to these methods, Albertsen et al., U.S. Pat. No. 5,432,068,describe a system of nuclear male sterility which includes: identifyinga gene which is critical to male fertility; silencing this native genewhich is critical to male fertility; removing the native promoter fromthe essential male fertility gene and replacing it with an induciblepromoter; inserting this genetically engineered gene back into theplant; and thus creating a plant that is male sterile because theinducible promoter is not “on” resulting in the male fertility gene notbeing transcribed. Fertility is restored by inducing, or turning “on”,the promoter, which in turn allows the gene that, confers male fertilityto be transcribed.

A. A tapetum-specific gene, RTS, a rice anther-specific gene is requiredfor male fertility and its promoter sequence directs tissue-specificgene expression in different plant species. Luo, Hong, et. al. (2006)Plant Molecular Biology. 62(3): 397-408(12). Introduction of adeacetylase gene under the control of a tapetum-specific promoter andwith the application of the chemical N-Ac-PPT. See internationalpublication number WO 01/29237.

B. Introduction of various stamen-specific promoters. Riceanther-specific promoters which are of particular utility in theproduction of transgenic male-sterile monocots and plants for restoringtheir fertility. See U.S. Pat. No. 5,639,948. See also internationalpublications WO 92/13956 and WO 92/13957.

C. Introduction of the barnase and the barstar genes. See Paul et al.,(1992) Plant Mol. Biol. 19:611-622.

For additional examples of nuclear male and female sterility systems andgenes, see also, U.S. Pat. Nos. 5,859,341, 6,297,426, 5,478,369,5,824,524, 5,850,014 and 6,265,640. See also Hanson, Maureen R., et.al., (2004) “Interactions of Mitochondrial and Nuclear Genes That AffectMale Gametophyte Development” Plant Cell. 16:S154-S169, all of which arehereby incorporated by reference.

5. Genes that Create a Site for Site Specific DNA Integration.

This includes the introduction of FRT sites that may be used in theFLP/FRT system and/or Lox sites that may be used in the Cre/Loxp system.For example, see Lyznik, et al., Site-Specific Recombination for GeneticEngineering in Plants, Plant Cell Rep (2003) 21:925-932 andinternational publication number WO 99/25821, which are herebyincorporated by reference. Other systems that may be used include theGin recombinase of phage Mu (Maeser et al., 1991; Vicki Chandler, TheMaize Handbook ch. 118 (Springer-Verlag 1994), the Pin recombinase of E.coli (Enomoto et al., 1983), and the R/RS system of the pSRi plasmid(Araki et al., 1992).

6. Genes that Affect Abiotic Stress Resistance.

Genes that affect abiotic stress resistance (including but not limitedto flowering, panicle/glume and seed development, enhancement ofnitrogen utilization efficiency, altered nitrogen responsiveness,drought resistance or tolerance, cold resistance or tolerance, and saltresistance or tolerance) and increased yield under stress. For example,see: Xiong, Lizhong, et al., (2003) “Disease Resistance and AbioticStress Tolerance in Rice Are Inversely Modulated by an AbscisicAcid—Inducible Mitogen-Activated Protein Kinase” The Plant Cell.15:745-759, where OsMAPK5 can positively regulate drought, salt, andcold tolerance and negatively modulate PR gene expression andbroad-spectrum disease resistance in rice; Chen, Fang, et. al., (2006)“The Rice 14-3-3 Gene Family and its Involvement in Responses to Bioticand Abiotic Stress” DNA Research 13(2):53-63, where at least four riceGF14 genes, GF14b, GF14c, GF14e and Gf14f, were differentially regulatedby salinity, drought, wounding and abscisic acid; internationalpublication number WO 00/73475 where water use efficiency is alteredthrough alteration of malate; U.S. Pat. Nos. 5,892,009, 5,965,705,5,929,305, 5,891,859, 6,417,428, 6,664,446, 6,717,034, 6,801,104 andInternational Publication Nos. WO 2000/060089, WO 2001/026459, WO2001/035725, WO 2001/034726, WO 2001/035727, WO 2001/036444, WO2001/036597, WO 2001/036598, WO 2002/015675, WO 2002/017430, WO2002/077185, WO 2002/079403, WO 2003/013227, WO 2003/013228, WO2003/014327, WO 2004/031349, WO 2004/076638, WO 98/09521 and WO 99/38977describing genes, including CBF genes and transcription factorseffective in mitigating the negative effects of freezing, high salinity,and drought on plants, as well as conferring other positive effects onplant phenotype; U.S. Publication No. 2004/0148654 and InternationalPublication No. WO 01/36596 where abscisic acid is altered in plantsresulting in improved plant phenotype such as increased yield and/orincreased tolerance to abiotic stress; International Publication Nos. WO2000/006341 and WO 04/090143, U.S. Publication No. 2004/0237147 and U.S.Pat. No. 6,992,237 where cytokinin expression is modified resulting inplants with increased stress tolerance, such as drought tolerance,and/or increased yield. Also see International Publication Nos. WO02/02776, WO 2003/052063, WO 01/64898, JP2002281975 and U.S. Pat. Nos.6,084,153, 6,177,275 and 6,107,547 (enhancement of nitrogen utilizationand altered nitrogen responsiveness). For ethylene alteration, see U.S.Publication Nos. 2004/0128719 and U.S 2003/0166197 and InternationalPublication No. WO 2000/32761. For plant transcription factors ortranscriptional regulators of abiotic stress, see e.g. U.S. PublicationNos. 2004/0098764 and 2004/0078852.

Other genes and transcription factors that affect plant growth andagronomic traits such as yield, flowering, plant growth and/or plantstructure, can be introduced or introgressed into plants, see e.g.International Publication Nos. WO 97/49811 (LHY), WO 98/56918 (ESD4), WO97/10339 WO 96/14414 (CON), WO 96/38560, WO 01/21822 (VRN1), WO 00/44918(VRN2), WO 99/49064 (GI), WO 00/46358 (FRI), WO 97/29123, WO 99/09174(D8 and Rht) and U.S. Pat. No. 6,573,430 (TFL), U.S. Pat. No. 6,713,663(FT), U.S. Pat. Nos. 6,794,560, 6,307,126 (GAI) and InternationalPublication Nos. WO 2004/076638 and WO 2004/031349 (transcriptionfactors).

Methods for Rice Transformation

Numerous methods for plant transformation have been developed, includingbiological and physical, plant transformation protocols. See, forexample, Miki et al., “Procedures for Introducing Foreign DNA intoPlants” in Methods in Plant Molecular Biology and Biotechnology, GlickB. R. and Thompson, J. E. Eds. (CRC Press, Inc., Boca Raton, 1993) pages67-88. In addition, expression vectors and in vitro culture methods forplant cell or tissue transformation and regeneration of plants areavailable. See, for example, Gruber et al., “Vectors for PlantTransformation” in Methods in Plant Molecular Biology and Biotechnology,Glick B. R. and Thompson, J. E. Eds. (CRC Press, Inc., Boca Raton, 1993)pages 89-119.

A. Agrobacterium-mediated Transformation—One method for introducing anexpression vector into plants is based on the natural transformationsystem of Agrobacterium. See, for example, Horsch et al., Science227:1229 (1985). A. tumefaciens and A. rhizogenes are plant pathogenicsoil bacteria which genetically transform plant cells. The Ti and Riplasmids of A. tumefaciens and A. rhizogenes, respectively, carry genesresponsible for genetic transformation of the plant. See, for example,Kado, C. I., Crit. Rev. Plant Sci. 10:1 (1991). Descriptions ofAgrobacterium vector systems and methods for Agrobacterium-mediated genetransfer are provided by Gruber et al., supra, Miki et al., supra, andMoloney et al., Plant Cell Reports 8:238 (1989). See also, U.S. Pat. No.5,591,616 issued Jan. 7, 1997.

B. Direct Gene Transfer—Despite the fact the host range forAgrobacterium-mediated transformation is broad, some major cereal cropspecies and gymnosperms have generally been recalcitrant to this mode ofgene transfer, even though some success has recently been achieved inrice and corn. Hiei et al., The Plant Journal 6:271-282 (1994) and U.S.Pat. No. 5,591,616 issued Jan. 7, 1997. Several methods of planttransformation collectively referred to as direct gene transfer havebeen developed as an alternative to Agrobacterium-mediatedtransformation.

A generally applicable method of plant transformation ismicroprojectile-mediated transformation wherein DNA is carried on thesurface of microprojectiles measuring 1 to 4 μm. The expression vectoris introduced into plant tissues with a biolistic device thataccelerates the microprojectiles to speeds of 300 to 600 m/s which issufficient to penetrate plant cell walls and membranes. Sanford et al.,Part. Sci. Technol. 5:27 (1987), Sanford, J. C., Trends Biotech. 6:299(1988), Klein et al., Bio/Technology 6:559-563 (1988), Sanford, J. C.,Physiol Plant 7:206 (1990), Klein et al., Biotechnology 10:268 (1992).In corn, several target tissues can be bombarded with DNA-coatedmicroprojectiles in order to produce transgenic plants, including, forexample, callus (Type I or Type II), immature embryos, and meristematictissue.

Another method for physical delivery of DNA to plants is sonication oftarget cells. Zhang et al., Bio/Technology 9:996 (1991). Additionally,liposome and spheroplast fusion have been used to introduce expressionvectors into plants. Deshayes et al., EMBO J., 4:2731 (1985), Christouet al., Proc Natl. Acad. Sci. U.S.A. 84:3962 (1987). Direct uptake ofDNA into protoplasts using CaCl₂ precipitation, polyvinyl alcohol orpoly-L-ornithine has also been reported. Hain et al., Mol. Gen. Genet.199:161 (1985) and Draper et al., Plant Cell Physiol. 23:451 (1982).Electroporation of protoplasts and whole cells and tissues have alsobeen described. Donn et al., In Abstracts of VIIth InternationalCongress on Plant Cell and Tissue Culture IAPTC, A2-38, p 53 (1990);D'Halluin et al., Plant Cell 4:1495-1505 (1992) and Spencer et al.,Plant Mol. Biol. 24:51-61 (1994).

Following transformation of rice target tissues, expression of theabove-described selectable marker genes allows for preferentialselection of transformed cells, tissues and/or plants, usingregeneration and selection methods now well known in the art.

Genetic Marker Profile Through SSR and First Generation Progeny

In addition to phenotypic observations, a plant can also be identifiedby its genotype. The genotype of a plant can be characterized through agenetic marker profile which can identify plants of the same variety ora related variety or be used to determine or validate a pedigree.Genetic marker profiles can be obtained by techniques such as IsozymeElectrophoresis, Restriction Fragment Length Polymorphisms (RFLPs),Randomly Amplified Polymorphic DNAs (RAPDs), Arbitrarily PrimedPolymerase Chain Reaction (AP-PCR), DNA Amplification Fingerprinting(DAF), Sequence Characterized Amplified Regions (SCARs), AmplifiedFragment Length Polymorphisms (AFLPs), Simple Sequence Repeats (SSRs)which are also referred to as Microsatellites, and Single NucleotidePolymorphisms (SNPs). For example, see Cregan et. al, “An IntegratedGenetic Linkage Map of the Soybean Genome” Crop Science 39:1464-1490(1999), and Berry et al., Assessing Probability of Ancestry Using SimpleSequence Repeat Profiles: Applications to Maize Inbred Lines and SoybeanVarieties” Genetics 165:331-342 (2003), each of which are incorporatedby reference herein in their entirety.

Particular markers used for these purposes are not limited to anyparticular set of markers, but are envisioned to include any type ofmarker and marker profile which provides a means of distinguishingvarieties. One method of comparison is to use only homozygous loci forhybrid rice HR120001.

Primers and PCR protocols for assaying these and other markers arewidely known in the art. In addition to being used for identification ofrice hybrid HR120001 and plant parts and plant cells of rice hybridHR120001, the genetic profile may be used to identify a rice plantproduced through the use of hybrid rice HR120001 or to verify a pedigreefor progeny plants produced through the use of hybrid rice HR120001. Thegenetic marker profile is also useful in breeding and developingbackcross conversions.

The present invention comprises a rice hybrid plant characterized bymolecular and physiological data obtained from the representative sampleof said hybrid deposited with the American Type Culture Collection(ATCC). Further provided by the invention is a rice hybrid plant formedby the combination of the disclosed rice hybrid plant or plant cell withanother rice plant or cell.

Means of performing genetic marker profiles using SSR polymorphisms arewell known in the art. SSRs are genetic markers based on polymorphismsin repeated nucleotide sequences, such as microsatellites. A markersystem based on SSRs can be highly informative in linkage analysisrelative to other marker systems in that multiple alleles may bepresent. Another advantage of this type of marker is that, through useof flanking primers, detection of SSRs can be achieved, for example, bythe polymerase chain reaction (PCR), thereby eliminating the need forlabor-intensive Southern hybridization. The PCR detection is done by useof two oligonucleotide primers flanking the polymorphic segment ofrepetitive DNA. Repeated cycles of heat denaturation of the DNA followedby annealing of the primers to their complementary sequences at lowtemperatures, and extension of the annealed primers with DNA polymerase,comprise the major part of the methodology.

Following amplification, markers can be scored by electrophoresis of theamplification products. Scoring of marker genotype is based on the sizeof the amplified fragment, which may be measured by the number of basepairs of the fragment. While variation in the primer used or inlaboratory procedures can affect the reported fragment size, relativevalues should remain constant regardless of the specific primer orlaboratory used. When comparing hybrids or varieties it is preferable ifall SSR profiles are performed in the same lab.

Primers used are publicly available and may be found in for example inU.S. Pat. Nos. 7,232,940, 7,217,003, 7,250,556, 7,214,851, 7,195,887 and7,192,774.

In addition, plants and plant parts substantially benefiting from theuse of rice hybrid HR120001 in their development, such as rice hybridHR120001 comprising a backcross conversion, transgene, or geneticsterility factor, may be identified by having a molecular marker profilewith a high percent identity to rice hybrid HR120001. Such a percentidentity might be 95%, 96%, 97%, 98%, 99%, 99.5% or 99.9% identical torice hybrid HR120001.

The SSR profile of rice hybrid HR120001 also can be used to identifyessentially derived varieties and other progeny varieties developed fromthe use of rice hybrid HR120001, as well as cells and other plant partsthereof. Such plants may be developed using the markers identified ininternational publication number WO 00/31964, U.S. Pat. No. 6,162,967and U.S. application Ser. No. 09/954,773. Progeny plants and plant partsproduced using rice hybrid HR120001 may be identified by having amolecular marker profile of at least 25%, 30%, 35%, 40%, 45%, 50%, 55%,60%, 65%, 70%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%,86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or99.5% genetic contribution from a rice hybrid or variety, as measured byeither percent identity or percent similarity. Such progeny may befurther characterized as being within a pedigree distance of rice hybridHR120001, such as within 1, 2, 3, 4 or 5 or fewer cross-pollinations toa rice plant other than rice hybrid HR120001 or a plant that has ricehybrid HR120001 as a progenitor. Unique molecular profiles may beidentified with other molecular tools such as SNPs and RFLPs.

While determining the SSR genetic marker profile of the plants describedsupra, several unique SSR profiles may also be identified which did notappear in either parent of such rice plant. Such unique SSR profiles mayarise during the breeding process from recombination or mutation. Acombination of several unique alleles provides a means of identifying aplant variety, an F₁ progeny produced from such variety, and progenyproduced from such rice plan.

Single or Multiple Gene Conversion

The foregoing methods for transformation would typically be used forproducing a transgenic hybrid or cultivar. The transgenic hybrid orcultivar could then be crossed, with another (non-transformed ortransformed) cultivar, in order to produce a new transgenic rice plant.Alternatively, a genetic trait which has been engineered into aparticular rice hybrid or cultivar using the foregoing transformationtechniques could be moved into another hybrid or cultivar usingtraditional backcrossing techniques that are well known in the plantbreeding arts. For example, a backcrossing approach could be used tomove an engineered trait from a public, non-elite rice plant into anelite rice plant, or from a rice plant containing a foreign gene in itsgenome into a rice plant which does not contain that gene. As usedherein, “crossing” can refer to a simple X by Y cross, or the process ofbackcrossing, depending on the context.

When the term rice plant is used in the context of the presentinvention, this also includes any single or multiple gene conversions ofthat rice hybrid or cultivar. The term single or multiple gene convertedplant as used herein refers to those rice plants which are developed bya plant breeding technique called backcrossing wherein essentially allof the desired morphological and physiological characteristics of acultivar are recovered in addition to the single gene transferred intothe cultivar via the backcrossing technique. Backcrossing methods can beused with the present invention to improve or introduce a characteristicinto the cultivar. The term backcrossing as used herein refers to therepeated crossing of a hybrid progeny back to one of the parental riceplants, the recurrent parent, for that cultivar, i.e., backcrossing 1,2, 3, 4, 5, 6, 7, 8, 9 or more times to the recurrent parent. Theparental rice plant which contributes the gene for the desiredcharacteristic is termed the nonrecurrent or donor parent. Thisterminology refers to the fact that the nonrecurrent parent is used onetime in the backcross protocol and therefore does not recur. Theparental rice plant to which the gene or genes from the nonrecurrentparent are transferred is known as the recurrent parent as it is usedfor several rounds in the backcrossing protocol (Jennings, P. R. et al.Rice Improvement (1979); Mackill D. On your mark, get, select. RiceToday, July-September pp 28-29 (2004); Fehr, W. R. et al. Principles ofCultivar Development—Theory and Technique pp. 261-286 (1987) andPohelman and Sleper (1994)).

In a typical backcross protocol, the original cultivar of interest(recurrent parent) is crossed to a second cultivar (nonrecurrent parent)that carries the single or multiple gene(s) of interest to betransferred. The resulting progeny from this cross are then crossedagain to the recurrent parent and the process is repeated until a riceplant is obtained wherein essentially all of the desired morphologicaland physiological characteristics of the recurrent parent are recoveredin the converted plant, in addition to the single or multipletransferred gene(s) from the nonrecurrent parent.

The selection of a suitable recurrent parent is an important step for asuccessful backcrossing procedure. The goal of a backcross protocol isto alter or substitute a single or multiple trait(s) orcharacteristic(s) in the original cultivar. To accomplish this, a singleor multiple gene(s) of the recurrent cultivar is modified or substitutedwith the desired gene(s) from the nonrecurrent parent, while retainingessentially all of the rest of the desired genetic, and therefore thedesired physiological and morphological, constitution of the originalcultivar. The choice of the particular nonrecurrent parent will dependon the purpose of the backcross; one of the major purposes is to addsome commercially desirable, agronomically important trait to the plant.The exact backcrossing protocol will depend on the characteristic(s) ortrait(s) being altered to determine an appropriate testing protocol.Although backcrossing methods are simplified when the characteristic(s)being transferred is the result of the action of a dominant allele(s), arecessive allele may also be transferred. In this instance it may benecessary to introduce a test of the progeny to determine if the desiredcharacteristic has been successfully transferred.

Many single or multiple gene traits have been identified that are notregularly selected for in the development of a new cultivar but that canbe improved by backcrossing techniques. Single or multiple gene traitsmay or may not be transgenic, examples of these traits include but arenot limited to, male sterility, waxy starch, herbicide resistance,resistance for bacterial, fungal, or viral disease, insect resistance,male fertility, enhanced nutritional quality, industrial usage, yieldstability and yield enhancement. These genes are generally inheritedthrough the nucleus. Some known exceptions to this are the genes formale sterility, some of which are inherited cytoplasmically, but stillact as single gene traits. Several of these single or multiple genetraits are described in U.S. Pat. Nos. 5,777,196; 5,948,957 and5,969,212, the disclosures of which are specifically hereby incorporatedby reference.

Introduction of a New Trait or Locus into Rice Hybrid HR120001

Rice hybrid HR120001 represents a new base genetic hybrid into which anew locus, loci or trait(s) may be introgressed. Direct transformationand backcrossing represent two important methods that can be used toaccomplish such an introgression. The term backcross conversion andsingle locus and multiple loci conversion are used interchangeably todesignate the product of a backcrossing program.

Backcross Conversions of Rice Hybrid HR120001

A backcross conversion of rice hybrid HR120001 occurs when DNA sequencesare introduced through backcrossing (Hallauer et al, 1988, “CornBreeding” Corn and Corn Improvements, No. 18, pp. 463-481), with ricehybrid HR120001 or either or both of the parental lines of rice hybridHR120001 are utilized as the recurrent parent. Both naturally occurringand transgenic DNA sequences may be introduced through backcrossingtechniques. A backcross conversion may produce a plant with a trait(s),locus or loci conversion in at least two or more backcrosses, includingat least 2 crosses, at least 3 crosses, at least 4 crosses, at least 5crosses and the like. Molecular marker assisted breeding or selectionmay be utilized to reduce the number of backcrosses necessary to achievethe backcross conversion. For example, see Openshaw, S. J. et al.,Marker-assisted Selection in Backcross Breeding. In: ProceedingsSymposium of the Analysis of Molecular Data, August 1994, Crop ScienceSociety of America, Corvallis, Oreg., where it is demonstrated that abackcross conversion can be made in as few as two backcrosses.

The complexity of the backcross conversion method depends on the type oftrait being transferred (single genes or closely linked genes as vs.unlinked genes), the level of expression of the trait, the type ofinheritance (cytoplasmic or nuclear) and the types of parents includedin the cross. It is understood by those of ordinary skill in the artthat for single gene traits that are relatively easy to classify, thebackcross method is effective and relatively easy to manage. (SeeHallauer et al. in Corn and Corn Improvement, Sprague and Dudley, ThirdEd. 1998). Desired traits that may be transferred through backcrossconversion include, but are not limited to, sterility (nuclear andcytoplasmic), fertility restoration, nutritional enhancements, droughttolerance, nitrogen utilization, altered fatty acid profile, lowphytate, industrial enhancements, disease resistance (bacterial, fungalor viral), insect resistance and herbicide resistance. In addition, anintrogression site itself, such as an FRT site, Lox site or other sitespecific integration site, may be inserted by backcrossing and utilizedfor direct insertion of one or more genes of interest into a specificplant variety. In some embodiments of the invention, the number of locithat may be backcrossed into rice hybrid HR120001 is at least 1, 2, 3,4, or 5 and/or no more than 6, 5, 4, 3, or 2. A single locus may containseveral transgenes, such as a transgene for disease resistance that, inthe same expression vector, also contains a transgene for herbicideresistance. The gene for herbicide resistance may be used as aselectable marker and/or as a phenotypic trait. A single locusconversion of a site specific integration system allows for theintegration of multiple genes at the converted loci.

Tissue Culture

Further reproduction of the hybrid can occur by tissue culture andregeneration. Tissue culture of various tissues of rice and regenerationof plants therefrom is well known and widely published. For example,reference may be had to Komatsuda, T. et al., Crop Sci. 31:333-337(1991); Stephens, P. A., et al., Theor. Appl. Genet. (1991) 82:633-635;Komatsuda, T. et al., Plant Cell, Tissue and Organ Culture, 28:103-113(1992); Dhir, S. et al., Plant Cell Reports (1992) 11:285-289; Pandey,P. et al., Japan J. Breed. 42:1-5 (1992); and Shetty, K., et al., PlantScience 81:245-251 (1992); as well as U.S. Pat. No. 5,024,944 issuedJun. 18, 1991 to Collins et al., and U.S. Pat. No. 5,008,200 issued Apr.16, 1991 to Ranch et al. Thus, another aspect of this invention is toprovide cells which upon growth and differentiation produce rice plantshaving the physiological and morphological characteristics of ricehybrid HR120001.

As used herein, the term “tissue culture” indicates a compositioncomprising isolated cells of the same or a different type or acollection of such cells organized into parts of a plant. Exemplarytypes of tissue cultures are protoplasts, calli, plant clumps, and plantcells that can generate tissue culture that are intact in plants orparts of plants, such as embryos, pollen, flowers, seeds, glumes,panicles, leaves, stems, roots, root tips, anthers, and the like. Meansfor preparing and maintaining plant tissue culture are well known in theart. By way of example, a tissue culture comprising organs has been usedto produce regenerated plants. U.S. Pat. Nos. 5,959,185; 5,973,234 and5,977,445 describe certain techniques, the disclosures of which areincorporated herein by reference.

As used herein, the term “plant” includes plant cells, plantprotoplasts, plant cells of tissue culture from which rice plants can beregenerated, plant calli, plant clumps, and plant cells that are intactin plants or parts of plants, such as pollen, flowers, embryos, ovules,seeds, panicles, glumes, leaves, stems, pistils, anthers and the like.Thus, another aspect of this invention is to provide for cells whichupon growth and differentiation produce a cultivar having essentiallyall of the physiological and morphological characteristics of HR120001.Genetic variants of rice hybrid HR120001 can also be obtained as aresult of the tissue culture process. Variants recovered by tissueculture of rice hybrid HR120001 are also the object of this invention.

The present invention contemplates a rice plant regenerated from atissue culture of the hybrid rice plant of the present invention. As iswell known in the art, tissue culture of rice can be used for the invitro regeneration of a rice plant. Tissue culture of various tissues ofrice and regeneration of plants therefrom is well known and widelypublished. For example, reference may be had to Chu, Q. R., et al.,(1999) “Use of bridging parents with high anther culturability toimprove plant regeneration and breeding value in rice”, RiceBiotechnology Quarterly 38:25-26; Chu, Q. R., et al., (1998), “A novelplant regeneration medium for rice anther culture of Southern U.S.crosses”, Rice Biotechnology Quarterly 35:15-16; Chu, Q. R., et al.,(1997), “A novel basal medium for embryogenic callus induction ofSouthern US crosses”, Rice Biotechnology Quarterly 32:19-20; and Oono,K., “Broadening the Genetic Variability By Tissue Culture Methods”, Jap.J. Breed. 33 (Suppl. 2), 306-307, illus. 1983. Thus, another aspect ofthis invention is to provide cells which upon growth and differentiationproduce rice plants having the physiological and morphologicalcharacteristics of hybrid HR120001.

Duncan, et al., Planta 165:322-332 (1985) reflects that 97% of theplants cultured that produced callus were capable of plant regeneration.Subsequent experiments with both cultivars and hybrids produced 91%regenerable callus that produced plants. In a further study in 1988,Songstad, et al., Plant Cell Reports 7:262-265 (1988), reports severalmedia additions that enhance regenerability of callus of two cultivars.Other published reports also indicated that “non-traditional” tissuesare capable of producing somatic embryogenesis and plant regeneration.K. P. Rao et al., Maize Genetics Cooperation Newsletter, 60:64-65(1986), refers to somatic embryogenesis from glume callus cultures andB. V. Conger, et al., Plant Cell Reports, 6:345-347 (1987) indicatessomatic embryogenesis from the tissue cultures of corn leaf segments.Thus, it is clear from the literature that the state of the art is suchthat these methods of obtaining plants are routinely used and have avery high rate of success.

Additional Breeding Methods

The utility of rice hybrid HR120001 also extends to crosses with otherspecies. Commonly, suitable species will be of the family Poaceae andespecially of the species sativa and glaberrima.

This invention also is directed to methods for producing a rice plant bycrossing a first parent rice plant with a second parent rice plantwherein the first or second parent rice plant is a rice plant of thehybrid HR120001. Further, both first and second parent rice plants cancome from the rice hybrid HR120001. Thus, any such methods using therice hybrid HR120001 are part of this invention: selfing, backcrosses,hybrid production, crosses to populations, and the like. All plantsproduced using rice hybrid HR120001 as a parent are within the scope ofthis invention, including those developed from varieties derived fromrice hybrid HR120001. Advantageously, the rice hybrid of the presentinvention could be used in crosses with other, different, rice plants toproduce the first generation (F₁) rice hybrid seeds and plants withsuperior characteristics. One or both parents of the hybrid of theinvention can also be used for transformation where exogenous genes areintroduced and expressed by one or both of the parents of the invention.Genetic variants created either through traditional breeding methodsusing one or both of the parents of rice hybrid HR120001 or throughtransformation of one or both of the parents of rice hybrid HR120001 byany of a number of protocols known to those of skill in the art areintended to be within the scope of this invention.

The following describes breeding methods that may be used with ricehybrid HR120001 or with one or both of the parents of rice hybridHR120001 in the development of further rice plants. One such embodimentis a method for developing an rice hybrid HR120001-derived progeny riceplant in a rice plant breeding program comprising: obtaining the riceplant, or a part thereof, of rice hybrid HR120001, utilizing said plantor plant part as a source of breeding material and selecting an ricehybrid HR120001 progeny plant with molecular markers in common with ricehybrid HR120001 and/or with morphological and/or physiologicalcharacteristics selected from the characteristics listed in Tables 1, 2,3, or 4. The same method may be used with one or both of the parents ofrice hybrid HR120001. Breeding steps that may be used in the rice plantbreeding program include pedigree breeding, back crossing, mutationbreeding, and recurrent selection. In conjunction with these steps,techniques such as RFLP-enhanced selection, genetic marker enhancedselection (for example SSR markers) and the making of double haploidsmay be utilized.

Another method involves producing a population of rice hybrid HR120001progeny rice plants, comprising crossing rice hybrid HR120001 withanother rice plant, thereby producing a population of rice plants,which, on average, derive 50% of their alleles from rice hybridHR120001. A plant of this population may be selected and repeatedlyselfed or sibbed with a rice cultivar resulting from these successivefilial generations. One embodiment of this invention is the ricecultivar produced by this method and that has obtained at least 50% ofits alleles from rice hybrid HR120001. The same method may be used withone or both of the parents of rice hybrid HR120001.

One of ordinary skill in the art of plant breeding would know how toevaluate the traits of two plant varieties to determine if there is nosignificant difference between the two traits expressed by thosevarieties. For example, see Fehr and Walt, Principles of CultivarDevelopment, p 261-286 (1987). Thus the invention includes rice hybridHR120001 progeny rice plants comprising a combination of at least tworice hybrid HR120001 traits selected from the group consisting of thoselisted in Tables 1, 2, 3, and 4 or the rice hybrid HR120001 combinationof traits listed in the Summary of the Invention, so that said progenyrice plant is not significantly different for said traits than ricehybrid HR120001. Using techniques described herein, molecular markersmay be used to identify said progeny plant as a rice hybrid HR120001progeny plant. Mean trait values may be used to determine whether traitdifferences are significant, and preferably the traits are measured onplants grown under the same environmental conditions. Once such avariety is developed its value is substantial since it is important toadvance the germplasm base as a whole in order to maintain or improvetraits such as yield, disease resistance, pest resistance, and plantperformance in extreme environmental conditions.

Progeny of rice hybrid HR120001 may also be characterized through theirfilial relationship with rice hybrid HR120001, as for example, beingwithin a certain number of breeding crosses of rice hybrid HR120001. Abreeding cross is a cross made to introduce new genetics into theprogeny, and is distinguished from a cross, such as a self or a sibcross, made to select among existing genetic alleles. The lower thenumber of breeding crosses in the pedigree, the closer the relationshipbetween rice hybrid HR120001 and its progeny. For example, progenyproduced by the methods described herein may be within 1, 2, 3, 4 or 5breeding crosses of rice hybrid HR120001.

Pedigree Breeding

Pedigree breeding starts with the crossing of two genotypes, such asrice hybrid HR120001 and another rice plant having one or more desirablecharacteristics that is lacking or which complements rice hybridHR120001. If the two original parents do not provide all the desiredcharacteristics, other sources can be included in the breedingpopulation. In the pedigree method, superior plants are selfed andselected in successive filial generations. In the succeeding filialgenerations the heterozygous condition gives way to homogeneousvarieties as a result of self-pollination and selection. Typically inthe pedigree method of breeding, five or more successive filialgenerations of selfing and selection is practiced: F₁ to F₂; F₂ to F₃;F₃ to F₄; F₄ to F₅, etc. After a sufficient amount of inbreeding,successive filial generations will serve to increase seed of thedeveloped variety. Preferably, the developed variety compriseshomozygous alleles at about 95% or more of its loci.

In addition to being used to create a backcross conversion, backcrossingcan also be used in combination with pedigree breeding. As discussedpreviously, backcrossing can be used to transfer one or morespecifically desirable traits from one variety, the donor parent, to adeveloped variety called the recurrent parent, which has overall goodagronomic characteristics yet lacks that desirable trait or traits.However, the same procedure can be used to move the progeny toward thegenotype of the recurrent parent but at the same time retain manycomponents of the non-recurrent parent by stopping the backcrossing atan early stage and proceeding with selfing and selection. For example, arice variety may be crossed with another rice variety to produce a firstgeneration progeny plant. The first generation progeny plant may then bebackcrossed to one of its parent varieties to create a BC1 or BC2.Progeny are selfed and selected so that the newly developed variety hasmany of the attributes of the recurrent parent and yet several of thedesired attributes of the non-recurrent parent. This approach leveragesthe value and strengths of the recurrent parent for use in new ricevarieties.

Therefore, an embodiment of this invention is a method of making abackcross conversion of rice hybrid HR120001, comprising the steps ofcrossing a plant of rice hybrid HR120001 with a donor plant comprising adesired trait, selecting an F₁ progeny plant comprising the desiredtrait, and backcrossing the selected F₁ progeny plant to a plant of ricehybrid HR120001. This method may further comprise the step of obtaininga molecular marker profile of rice hybrid HR120001 and using themolecular marker profile to select for a progeny plant with the desiredtrait and the molecular marker profile of rice hybrid HR120001. In oneembodiment the desired trait is a mutant gene or transgene present inthe donor parent.

Recurrent Selection and Mass Selection

Recurrent selection is a method used in a plant breeding program toimprove a population of plants. Rice hybrid HR120001 is suitable for usein a recurrent selection program. The method entails individual plantscross pollinating with each other to form progeny. The progeny are grownand the superior progeny selected by any number of selection methods,which include individual plant, half-sib progeny, full-sib progeny andselfed progeny. The selected progeny are cross pollinated with eachother to form progeny for another population. This population is plantedand again superior plants are selected to cross pollinate with eachother. Recurrent selection is a cyclical process and therefore can berepeated as many times as desired. The objective of recurrent selectionis to improve the traits of a population. The improved population canthen be used as a source of breeding material to obtain new varietiesfor commercial or breeding use, including the production of a syntheticcultivar. A synthetic cultivar is the resultant progeny formed by theintercrossing of several selected varieties.

Mass selection is a useful technique when used in conjunction withmolecular marker enhanced selection. In mass selection seeds fromindividuals are selected based on phenotype or genotype. These selectedseeds are then bulked and used to grow the next generation. Bulkselection requires growing a population of plants in a bulk plot,allowing the plants to self-pollinate, harvesting the seed in bulk andthen using a sample of the seed harvested in bulk to plant the nextgeneration. Also, instead of self pollination, directed pollinationcould be used as part of the breeding program.

Mutation Breeding

Mutation breeding is another method of introducing new traits into ricehybrid HR120001. Mutations that occur spontaneously or are artificiallyinduced can be useful sources of variability for a plant breeder. Thegoal of artificial mutagenesis is to increase the rate of mutation for adesired characteristic. Mutation rates can be increased by manydifferent means including temperature, long-term seed storage, tissueculture conditions, radiation; such as X-rays, Gamma rays (e.g. cobalt60 or cesium 137), neutrons, (product of nuclear fission by uranium 235in an atomic reactor), Beta radiation (emitted from radioisotopes suchas phosphorus 32 or carbon 14), or ultraviolet radiation (preferablyfrom 2500 to 2900 nm), or chemical mutagens (such as base analogues(5-bromo-uracil), related compounds (8-ethoxy caffeine), antibiotics(streptonigrin), alkylating agents (sulfur mustards, nitrogen mustards,epoxides, ethylenamines, sulfates, sulfonates, sulfones, lactones),azide, hydroxylamine, nitrous acid, or acridines. Once a desired traitis observed through mutagenesis the trait may then be incorporated intoexisting germplasm by traditional breeding techniques. Details ofmutation breeding can be found in “Principles of Cultivar Development”Fehr, 1993 Macmillan Publishing Company. In addition, mutations createdin other rice plants may be used to produce a backcross conversion ofrice hybrid HR120001 that comprises such mutation.

Breeding with Molecular Markers

Molecular markers may be used in plant breeding methods utilizing ricehybrid HR120001.

Isozyme Electrophoresis and RFLPs have been widely used to determinegenetic composition. See for example, Dinka, S. J., et al. (2007)“Predicting the size of the progeny mapping population required topositionally clone a gene” Genetics. 176(4):2035-54; Gonzalez, C., etal. (2007) “Molecular and pathogenic characterization of new Xanthomonasoryzae strains from West Africa” Mol. Plant. Microbe Interact.20(5):534-546; Jin, H., et al. (2006) “Molecular and cytogeniccharacterization of an Oryza officinalis-O. sativa chromosome 4 additionline and its progenies” Plant Mol. Biol. 62(4-5):769-777; Pan, G., etal. (2006) “Map-based cloning of a novel rice cytochrome P450 geneCYP81A6 that confers resistance to two different classes of herbicides”Plant Mol. Biol. 61(6):933-943.; Huang, W., et al. (2007) “RFLP analysisfor mitochondrial genome of CMS-rice” Journal of Genetics and Genomics.33(4):330-338; Yan, C. J., et al. (2007) “Identification andcharacterization of a major QTL responsible for erect panicle trait injaponica rice (Oryza sativa L.)” Theor. Appl. Genetics.DOI:10.1007/s00122-007-0635-9; and I. K. Vasil (ed.) DNA-based markersin plants. Kluwer Academic Press Dordrecht, the Netherlands.

SSR technology is currently the most efficient and practical markertechnology; more marker loci can be routinely used and more alleles permarker locus can be found using SSRs in comparison to RFLPs. Gealy,David, et al. (2005) “Insights into the Parentage of Rice/red RiceCrosses Using SSR Analysis of US Rice Cultivars and Red RicePopulations”. Rice Technical Working Group Meeting Proceedings. Abstractp. 179.; Lawson, Mark J., et al. (2006) “Distinct Patterns of SSRDistribution in the Arabidopsis thaliana and rice genomes” GenomeBiology. 7:R14; Nagaraju, J., et al., (2002) “Genetic Analysis ofTraditional and Evolved Basmati and Non-Basmati Rice Varieties by UsingFluorescence-based ISSR-PCR and SSR Markers” Proc. Nat. Acad. Sci. USA.99(9):5836-5841; and Lu, Hong, et al. (2005) “Population Structure andBreeding Patterns of 145 US Rice Cultivars Based on SSR Marker Analysis”Crop Science. 45:66-76. Single Nucleotide Polymorphisms may also be usedto identify the unique genetic composition of the invention and progenyvarieties retaining that unique genetic composition. Various molecularmarker techniques may be used in combination to enhance overallresolution.

Rice DNA molecular marker linkage maps have been rapidly constructed andwidely implemented in genetic studies such as in Zhu, J. H., et al.(1999) “Toward rice genome scanning by map-based AFLP fingerprinting”Mol. Gene Genetics. 261(1):184-195; Cheng, Z., et al (2001) “Toward acytological characterization of the rice genome” Genome Research.11(12):2133-2141; Ahn, S., et al. (1993) “Comparative linkage maps ofthe rice and maize genomes” Proc. Natl. Acad. Sci. USA.90(17):7980-7984; and Kao, F. I., et al. (2006) “An integrated map ofOryza sativa L. chromosome 5” Theor. Appl. Genet. 112(5):891-902.Sequences and PCR conditions of SSR Loci in rice as well as the mostcurrent genetic map may be found in RiceBLAST and the TIGR Rice GenomeAnnotation on the World Wide Web.

One use of molecular markers is Quantitative Trait Loci (QTL) mapping.QTL mapping is the use of markers, which are known to be closely linkedto alleles that have measurable effects on a quantitative trait.Selection in the breeding process is based upon the accumulation ofmarkers linked to the positive effecting alleles and/or the eliminationof the markers linked to the negative effecting alleles from the plant'sgenome.

Molecular markers can also be used during the breeding process for theselection of qualitative traits. For example, markers closely linked toalleles or markers containing sequences within the actual alleles ofinterest can be used to select plants that contain the alleles ofinterest during a backcrossing breeding program. The markers can also beused to select for the genome of the recurrent parent and against thegenome of the donor parent. Using this procedure can minimize the amountof genome from the donor parent that remains in the selected plants. Itcan also be used to reduce the number of crosses back to the recurrentparent needed in a backcrossing program. The use of molecular markers inthe selection process is often called genetic marker enhanced selection.Molecular markers may also be used to identify and exclude certainsources of germplasm as parental varieties or ancestors of a plant byproviding a means of tracking genetic profiles through crosses.

Production of Double Haploids

The production of double haploids can also be used for the developmentof plants with a homozygous phenotype in the breeding program. Forexample, a rice plant for which rice hybrid HR120001 is a parent can beused to produce double haploid plants. Double haploids are produced bythe doubling of a set of chromosomes (1 N) from a heterozygous plant toproduce a completely homozygous individual. For example, see Wan et al.,“Efficient Production of Doubled Haploid Plants Through ColchicineTreatment of Anther-Derived Maize Callus”, Theoretical and AppliedGenetics, 77:889-892, 1989 and U.S. Pat. No. 7,135,615.

Methods for obtaining haploid plants are also disclosed in Kobayashi, M.et al., Journ. of Heredity 71(1):9-14, 1980, Pollacsek, M., Agronomie(Paris) 12(3):247-251, 1992; Cho-Un-Haing et al., Journ. of Plant Biol.,1996, 39(3):185-188; Verdoodt, L., et al., February 1998, 96(2):294-300;Genetic Manipulation in Plant Breeding, Proceedings InternationalSymposium Organized by EUCARPIA, Sep. 8-13, 1985, Berlin, Germany;Thomas, W J K, et al. (2003) “Doubled haploids in breeding” in DoubledHaploid Production in Crop Plants. Maluszynski, M., et al. (Eds.)Dordrecht, the Netherland Kluwer Academic Publishers. pp. 337-349.

Descriptions of other breeding methods that are commonly used fordifferent traits and crops can be found in one of several referencebooks (e.g., Allard, 1960; Simmonds, 1979; Sneep et al., 1979; Fehr,1987).

The seed of rice hybrid HR120001, the plant produced from the hybridseed, the hybrid rice plant produced from the crossing of the hybrid,and various parts of the hybrid rice plant and transgenic versions ofthe foregoing, can be utilized for human food, livestock feed, and as araw material in industry.

Tables

The following tables present data on the traits and characteristics ofrice hybrid HR120001 as compared with Chemere, a commonly grown ricevariety in the U.S. The data was collected from multiple locations andrepeated trials. In the following tables, probability figures indicatethe probability associated with a paired Student's t-Test used todetermine whether two samples are likely to have come from the same twounderlying populations that have the same mean. The N.S. notation meansthat there is no significant difference between the means of the twosamples.

In Table 2, column 2 shows the yield in kilograms per hectare, column 3shows the plant height in centimeters, column 4 shows the number of daysto 50% flowering, column 5 shows the percent lodging, column 6 shows thetotal milling percent, and column 7 shows the whole milling percent. Thenumber of different locations over which the data was collected is shownin row 4. The data compares yield, plant height, maturity, lodging, andmilling yields of Chemere versus rice hybrid HR120001.

As shown in Table 2, rice hybrid HR120001 has significantly higheryield, plant height, and flowers earlier than the cultivar Chemere. Yet,Chemere has significantly higher total milling and whole millingpercentage than rice hybrid HR120001. Both the rice hybrid HR120001 andthe cultivar Chemere have similar lodging percent.

TABLE 2 Plant Days Total Whole Yield Height to 50% Lodging MillingMilling (kg/ha) (cm) Flowering % % % HR120001 9501 101 72 1.0 72.4 62.0Cheniere 8004 96 83 4.7 73.8 67.0 Locations 14 11 7 14 13 13 Difference1497 5 −11 −3.7 −1.4 −5.0 Probability .001 .001 .001 N.S. .001 .001

In Table 3, column 2 shows the amylose percentage, column 3 shows thealkali spreading value (ASV), column 4 shows the milled grain length inmillimeters, column 5 shows the milled grain width in millimeters,column 6 shows the milled grain length to width ratio, column 7 showsthe milled grain thickness in millimeters, and column 8 shows the milledgrain chalk percent. The number of different locations over which thedata was collected is shown in row 4. The data compares the basicquality characteristics of rice hybrid HR120001 and Chemere.

Unexpectedly, rice hybrid HR120001 had significantly lower amylose,gelatinization temperature (ASV), and higher chalk when compared toChemere. Yet, milled grain length, milled grain width, milled grainlength to width ratio, and milled grain thickness are the same for bothChemere and rice hybrid HR120001.

TABLE 3 Thick- Amylose Length Width L/W ness Chalk % ASV (mm) (mm) Ratio(mm) % HR120001 24 3.8 6.9 2.1 3.3 1.8 9 Cheniere 25 4.5 6.9 2.1 3.3 1.82 Locations 12 12 12 12 12 18 12 Difference −1 −0.7 0.0 0.0 0.0 0.0 7Probability .05 .001 N.S. N.S. N.S. N.S .01

In the next 2 tables, grain quality characteristics of rice hybridHR120001 are compared to Chemere.

In Table 4, column 2 shows the peak viscosity expressed in RapidVisco-Analyser units (RVU), column 3 shows the peak time in minutes,column 4 shows the trough in RVU, column 5 shows the trough time inminutes, column 6 shows the paste temperature in degrees Celsius, andcolumn 7 shows the paste time in minutes. In table 5, column 2 shows thefinal viscosity in RVU, column 3 shows the breakdown in RVU, column 4shows the setback in RVU, and column 5 shows the consistency of thestarch in RVU, column 6 shows the whiteness, and in column 7 shows thetransparency. The whiteness and transparency are expressed in lightreflectance and transparency units, respectively, as measured by theSakate Milling Degree Meter.

Rice hybrid HR120001 has a higher peak viscosity, peak time, trough,final viscosity, breakdown, setback, and consistency than Chemere. Yetthe trough time was the same for both, HR120001 and Chemere. For pastetemperature, paste time, whiteness, and transparency rice hybridHR120001 has lower values than Chemere.

TABLE 4 Paste Paste Peak Peak Trough Temper- Time Viscosity Time TroughTime ature (min- (RVU) (minutes) (RVU) (minutes) (° C.) utes) HR120001211.1 5.8 128.7 8.2 87.2 4.1 Cheniere 146.8 5.6  85.1 8.2 89.1 4.3Difference  64.3 0.2  43.6 0.0 −1.9 −0.2

TABLE 5 White- Trans- ness parency Final Break Set Consis- (light (lightViscosity Down Back tency reflec- trans- (RVU) (RVU) (RVU) (RVU) tance)mission) HR120001 266.4 82.6 55.3 137.8 50.1 2.6 Cheniere 200.4 61.753.5 115.3 51.3 3.2 Difference  66.0 20.9  1.8  22.5 −1.2 −0.6

EXAMPLES

The following examples further describe the materials and methods usedin carrying out the invention and the subsequent results. It should beunderstood that these Examples, while indicating preferred embodimentsof the invention, are given by way of illustration only. From the abovediscussion and these Examples, one skilled in the art can ascertain theessential characteristics of this invention, and without departing fromthe spirit and scope thereof, can make various changes and modificationsof the invention to adapt it to various usages and conditions.

Example 1 Growing a Blend of Rice Seed of Rice Hybrid HR120001 withAnother Rice Hybrid, Inbred or Variety

Another method of the present invention is producing a blend of riceseeds by planting the rice seed of rice hybrid HR120001 with rice seedof one, two, three, four, five or more of another rice variety, riceinbred or rice hybrid. The seed of rice hybrid HR120001 is present in arange of about 1% to about 95%. The blend of rice seeds are then grownto produce rice plants. The seeds of rice hybrid HR120001 contained inthe blend and the seeds of one, two, three, four, five or more ofanother rice variety, rice inbred or rice hybrid also contained in theblend are grown in the same field and then harvested together.

Example 2 Preparing a Blend of Rice Seed Using Rice Hybrid HR120001

Another method of the present invention is producing a blend of riceseed using the rice seed of rice hybrid HR120001. The blend consists ofproviding for a first quantity of rice seed of rice hybrid HR120001,providing for a second, third, fourth, fifth or higher quantity of riceseed of another rice variety, rice hybrid or rice inbred and mixing allquantities of rice seed to produce a blend of rice seed containinghybrid rice HR120001 in a range of about 1% to about 95% and rice seedof one, two, three, four, five or more of another rice hybrid, riceinbred or rice variety.

The use of the terms “a,” “an,” and “the,” and similar referents in thecontext of describing the invention (especially in the context of thefollowing claims) are to be construed to cover both the singular and theplural, unless otherwise indicated herein or clearly contradicted bycontext. The terms “comprising,” “having,” “including,” and “containing”are to be construed as open-ended terms (i.e., meaning “including, butnot limited to,”) unless otherwise noted. Recitation of ranges of valuesherein are merely intended to serve as a shorthand method of referringindividually to each separate value falling within the range, unlessotherwise indicated herein, and each separate value is incorporated intothe specification as if it were individually recited herein. Forexample, if the range 10-15 is disclosed, then 11, 12, 13, and 14 arealso disclosed. All methods described herein can be performed in anysuitable order unless otherwise indicated herein or otherwise clearlycontradicted by context. The use of any and all examples, or exemplarylanguage (e.g., “such as”) provided herein, is intended merely to betterilluminate the invention and does not pose a limitation on the scope ofthe invention unless otherwise claimed. No language in the specificationshould be construed as indicating any non claimed element as essentialto the practice of the invention.

Deposit Information

An ATCC deposit of the rice seed (PTA-13290) of this invention accordingto the Budapest Treaty on the International Recognition of the Depositof Microorganisms for the Purposes of Patent Procedure, is owned byRiceTec, Inc. and is maintained by RiceTec, Inc., 1925 FM 2917, Alvin,Tex. 77511. Access to this deposit will be available during the pendencyof this application to persons determined by the Commissioner of Patentsand Trademarks to be entitled thereto under 37 CFR 1.14 and 35 USC 122.Upon allowance of any claims in this application, all restrictions onthe availability to the public of the inbred line will be irrevocablyremoved by affording access to a deposit of at least 2,500 seeds of thesame inbred line with the American Type Culture Collection, Manassas,Va.

While a number of exemplary aspects and embodiments have been discussedabove, those of skill in the art will recognize certain modifications,permutations, additions and sub-combinations thereof. It is thereforeintended that the following appended claims and claims hereafterintroduced are interpreted to include all such modifications,permutations, additions and sub-combinations as are within their truespirit and scope.

What is claimed is:
 1. A hybrid rice seed designated HR120001, wherein arepresentative sample of seed of said hybrid rice was deposited underATCC Accession No. PTA-13290.
 2. A rice plant, or a part thereof,produced by growing the seed of claim
 1. 3. Pollen or an ovule of theplant of claim
 2. 4. A rice plant or a part thereof having all of thephysiological and morphological characteristics of the rice plant ofclaim
 2. 5. A tissue culture produced from protoplasts or cells from therice plant of claim 2, wherein said cells or protoplasts of the tissueculture are produced from a plant part selected from the groupconsisting of leaves, pollen, embryos, cotyledon, hypocotyl,meristematic cells, roots, root tips, pistils, anthers, flowers, stems,glumes and panicles.
 6. A protoplast produced from the plant of claim 2.7. A rice plant regenerated from the tissue culture of claim 5, whereinthe plant has all the morphological and physiological characteristics ofrice hybrid HR120001.
 8. A method for producing rice seed, wherein themethod comprises crossing the plant of claim 2 with a different riceplant and harvesting the resultant hybrid rice seed.
 9. A hybrid riceseed produced by the method of claim
 8. 10. A method of producing anherbicide resistant rice plant, wherein the method comprisestransforming the rice plant of claim 2 with a transgene, wherein thetransgene confers resistance to an herbicide selected from the groupconsisting of imidazolinone, cyclohexanedione, sulfonylurea, glyphosate,glufosinate, phenoxy proprionic acid, L-phosphinothricin, triazine andbenzonitrile.
 11. An herbicide resistant rice plant produced by themethod of claim
 10. 12. A method of producing a pest or insect resistantrice plant, wherein the method comprises transforming the rice plant ofclaim 2 with a transgene that confers insect resistance.
 13. A pest orinsect resistant rice plant produced by the method of claim
 12. 14. Therice plant of claim 13, wherein the transgene encodes a Bacillusthuringiensis endotoxin.
 15. A method of producing a disease resistantrice plant, wherein the method comprises transforming the rice plant ofclaim 2 with a transgene that confers disease resistance.
 16. A diseaseresistant rice plant produced by the method of claim
 15. 17. A method ofproducing a rice plant with modified fatty acid metabolism or modifiedcarbohydrate metabolism, wherein the method comprises transforming therice plant of claim 2 with a transgene encoding a protein selected fromthe group consisting of fructosyltransferase, levansucrase,alpha-amylase, invertase and starch branching enzyme or DNA encoding anantisense of stearyl-ACP desaturase.
 18. A rice plant having modifiedfatty acid metabolism or modified carbohydrate metabolism produced bythe method of claim
 17. 19. A method of growing a blend of rice seed,wherein the method comprises: (a) planting a blend comprising a firstquantity of rice seed of claim 1 mixed with a second quantity of riceseed of another rice variety, rice hybrid or rice inbred; (b) growingsaid seeds to produce rice plants; and (c) harvesting seeds from saidrice plants.
 20. The method of claim 19, wherein said blend is comprisedof seeds from a third, fourth or fifth rice variety, rice hybrid or riceinbred.
 21. The method of claim 19, wherein said blend is comprised ofabout 1% to about 95% of rice hybrid HR120001 seed.
 22. A method ofproducing a blend of rice seed, wherein the method comprises: (a)providing a first quantity of rice seed of claim 1; (b) providing asecond quantity of rice seed of another rice variety, rice inbred orrice hybrid; and (c) producing a blend comprised of mixing said firstquantity of rice seed with said second quantity of rice seed.
 23. Themethod of claim 22 wherein said blend consists of seeds from a third,fourth or fifth rice variety, rice inbred, or rice hybrid.
 24. Themethod of claim 22, wherein said blend is comprised of about 1% to about95% of rice hybrid HR120001 seed.